Academic literature on the topic 'CRISPR/Cas 9'

“CRISPR" is short and used for "CRISPR-Cas9. CRISPR stands for clustered regularly interspaced short palindromic repeats. CRISPRs are specialized stretches of DNA. The protein Cas9 (or "CRISPR-associated") is an enzyme that acts like a pair of molecular scissors, capable of cutting strands of DNA and can be used in conjunction with engineered CRISPR sequences to hunt down codes and slice into them like a molecular scalpel, allowing geneticists to cut out a target gene, either to remove it or replace it with a new sequence. Therefore it is a simple and powerful tool for editing genomes to easily alter DNA sequences and amend gene function. In 1987, The CRISPR locus was first identified in Escherichia coli and discovered when a genetic structure containing 5 highly homologous repeats of 29 nucleotides separated by 32-nucleotide spacers (Ishino Y 1987).

CRISPR-Cas9 technology is reshaping the way scientists conduct research in genetic engineering. It is predicted to revolutionise not only the fields of medicine, biology, agriculture and industry but, much like all revolutionary technologies of the past, the way humans live. Given the anticipated and already seen benefits of CRISPR-Cas 9 in different areas of human life, this new technology may be defined as a true breakthrough scientific discovery. The article presents several challenges connected with various dimensions of the CRISPR-Cas 9 patent landscape. The central argument is that today the biggest challenge is finding a intermediary way that ensures a balance between providing sufficient openness for the further progress of basic research in CRISPR-Cas 9 such as ‘niche’ areas of the latest genetic engineering and adequate intellectual property rights to incentivise its commercialisation and application. The article contends the endeavours by academic scientific institutions to arrive at short-term benefits of the new CRISPR-Cas 9 technology do not constitute such an intermediary way, especially when the CRISPR-Cas 9 patent landscape is viewed as part of a series of controversial bioethical discussions that have been underway for over 40 years.

Organoids are three-dimensional (3D) structures derived from adult or embryonic stem cells that maintain many structural and functional features of their respective organ. Recently, genome editing based on the bacterial defense mechanism CRISPR/Cas9 has emerged as an easily applicable and reliable laboratory tool. Combining organoids and CRISPR/Cas9 creates exciting new opportunities to study organ development and human disease in vitro. The potential applications of CRISPR in organoids are only beginning to be explored.

Objetiva-se analisar as utilizações do sistema Crispr/Cas-9 no melhoramento vegetal, por meio de revisão sistemática de literatura realizada na base de dados Scopus. Elaborou-se um protocolo de pesquisa para selecionar os artigos a partir de critérios de exclusão e inclusão. Os dados desta pesquisa indicam a maior concentração de trabalhos no continente asiático, tendo a China se apresentando como principal expoente em pesquisas na área, com um foco muito grande no melhoramento de Oriza sativa e Solanum lycopersicum. Esses estudos na maioria das vezes buscam o aumento na produtividade e a tolerância a estresse abiótico e biótico isso por que há uma crescente busca por melhoria da qualidade dos grãos e melhoria da resistência a estresses diante do aquecimento climático. Palavras-chaves: Biotecnologia. Edição genética em plantas. Espécies vegetais.

Abstract The rapid development of CRISPR–Cas technologies brought a personalized and targeted treatment of genetic disorders into closer reach. To render CRISPR-based therapies precise and safe, strategies to confine the activity of Cas(9) to selected cells and tissues are highly desired. Here, we developed a cell type-specific Cas-ON switch based on miRNA-regulated expression of anti-CRISPR (Acr) proteins. We inserted target sites for miR-122 or miR-1, which are abundant specifically in liver and cardiac muscle cells, respectively, into the 3′UTR of Acr transgenes. Co-expressing these with Cas9 and sgRNAs resulted in Acr knockdown and released Cas9 activity solely in hepatocytes or cardiomyocytes, while Cas9 was efficiently inhibited in off-target cells. We demonstrate control of genome editing and gene activation using a miR-dependent AcrIIA4 in combination with different Streptococcus pyogenes (Spy)Cas9 variants (full-length Cas9, split-Cas9, dCas9-VP64). Finally, to showcase its modularity, we adapted our Cas-ON system to the smaller and more target-specific Neisseria meningitidis (Nme)Cas9 orthologue and its cognate inhibitors AcrIIC1 and AcrIIC3. Our Cas-ON switch should facilitate cell-specific activity of any CRISPR–Cas orthologue, for which a potent anti-CRISPR protein is known.

Background: Certainly, the ultimate aim of Islamic law is to “protect human life” either through mitigation of hardship or recognition of public interests reckons biomedical innovations allowable where-if cling by ethical, moral and legal principles. Assertively, if–CRISPR Cas- 9 genome editing–methods based on the guided principles of Islamic law and jurisprudence, as “harm has to be redressed” can be justified keeping in view the human dignity, honor and prestige. Hence, newer technologies can be adopted because “necessity renders prohibited things as permissible” with certain caveats. Arguably those who consider it as an evil must think over that “in the presence of two evils, the one whose injury is greater is avoided by the commission of the lesser”. Conclusion: Therefore if Cas-9 based method leaning towards evils, even then it can be acceptable in case where an atypical germ-line sequence can affect the next generation, which is indeed a great evil, and “the lesser of evils is preferred over the greater one” renders it permissible with a view it might enhance human health and living standard. Conversely, curing a minor disease if causing another equal infirmity or greater should be rendered forbidden as “harm cannot be removed by harm”, then, “a greater harm can be removed by a lesser one” germ-line editing/alteration in severe cases will be allowed on the basis of necessity. Bangladesh Journal of Medical Science Vol.18(1) 2019 p.7-13

ABSTRACTClustered regularly interspaced short palindromic repeats (CRISPR) in combination with associated sequences (cas) constitute the CRISPR-Cas immune system, which uptakes DNA from invasive genetic elements as novel “spacers” that provide a genetic record of immunization events. We investigated the potential of CRISPR-based genotyping ofLactobacillus buchneri, a species relevant for commercial silage, bioethanol, and vegetable fermentations. Upon investigating the occurrence and diversity of CRISPR-Cas systems inLactobacillus buchnerigenomes, we observed a ubiquitous occurrence of CRISPR arrays containing a 36-nucleotide (nt) type II-A CRISPR locus adjacent to fourcasgenes, including the universalcas1andcas2genes and the type II signature genecas9. Comparative analysis of CRISPR spacer content in 26L. buchneripickle fermentation isolates associated with spoilage revealed 10 unique locus genotypes that contained between 9 and 29 variable spacers. We observed a set of conserved spacers at the ancestral end, reflecting a common origin, as well as leader-end polymorphisms, reflecting recent divergence. Some of these spacers showed perfect identity with phage sequences, and many spacers showed homology toLactobacillusplasmid sequences. Following a comparative analysis of sequences immediately flanking protospacers that matched CRISPR spacers, we identified a novel putative protospacer-adjacent motif (PAM), 5′-AAAA-3′. Overall, these findings suggest that type II-A CRISPR-Cas systems are valuable for genotyping ofL. buchneri.

Atherosclerosis represents one of the major causes of death globally. The high mortality rates and limitations of current therapeutic modalities have urged researchers to explore potential alternative therapies. The clustered regularly interspaced short palindromic repeats-associated protein 9 (CRISPR/Cas9) system is commonly deployed for investigating the genetic aspects of Atherosclerosis. Besides, advances in CRISPR/Cas system has led to extensive options for researchers to study the pathogenesis of this disease. The recent discovery of Cas9 variants, such as dCas9, Cas9n, and xCas9 have been established for various applications, including single base editing, regulation of gene expression, live-cell imaging, epigenetic modification, and genome landscaping. Meanwhile, other Cas proteins, such as Cas12 and Cas13, are gaining popularity for their applications in nucleic acid detection and single-base DNA/RNA modifications. To date, many studies have utilized the CRISPR/Cas9 system to generate disease models of atherosclerosis and identify potential molecular targets that are associated with atherosclerosis. These studies provided proof-of-concept evidence which have established the feasibility of implementing the CRISPR/Cas system in correcting disease-causing alleles. The CRISPR/Cas system holds great potential to be developed as a targeted treatment for patients who are suffering from atherosclerosis. This review highlights the advances in CRISPR/Cas systems and their applications in establishing pathogenetic and therapeutic role of specific genes in atherosclerosis.

Model organisms and gene-editing strategies are fundamental to address a variety of scientific questions from basic science to biomedical research. Here, we reinforced the use of two powerful tools, the experimental system Caenorhabditis elegans and the CRISPR/Cas gene-editing technology, to model cancer-related mutations and investigate cisplatin-based chemoresistance. We have established a model to study BAP1 cancer predisposition syndrome-related mutations in the BAP1 ortholog ubh-4. By exploring distinct ubh-4 alleles, we have discovered a synthetic interaction between ubh-4 and rpn-9, an essential regulatory subunit involved in proteasome assembly. Moreover, we suggest a cooperating role between these genes in the ubiquitin-mediated proteostasis at meiotic prophase. In addition, we have exploited C. elegans to study the toxicity of cisplatin-based therapies in different ways. First, by studying the impact of glucose and lipid metabolism on cisplatin toxicity. Then, we have described the harmful effect of cisplatin in mitochondrial functions. Finally, we have established a method to investigate the cisplatin-induced neurotoxicity by using an automated worm tracking system and discovered a protective role of dopamine.
Los organismos modelo y las estrategias de edición genética son fundamentales para desentrañar incógnitas en ciencias de la vida, desde la investigación básica hasta investigación aplicada a la biomedicina. En este estudio, reafirmamos la importancia del uso de dos potentes herramientas, el sistema experimental Caenorhabditis elegans y la tecnología de edición genética CRISPR/Cas, para modelar mutaciones relacionadas con cáncer e investigar la quimiorresistencia al cisplatino. Hemos modelado mutaciones asociadas al síndrome de predisposición tumoral BAP1, en ubh-4/BAP1. Explorando el efecto de distintos alelos mutantes de ubh-4, hemos descubierto una interacción sintética entre ubh-4 y rpn-9, el cual codifica para una subunidad reguladora esencial para el ensamblaje del proteasoma. Además, proponemos que la cooperación funcional de dichos genes está implicada en la degradación de proteínas mediada por el sistema ubiquitina-proteasoma durante la profase meiótica. También hemos investigado la respuesta generada por la terapia con cisplatino en C. elegans. Por una parte, hemos demostrado que la toxicidad inducida por el cisplatino puede modularse alterando el metabolismo glucídico y lipídico. Por otro lado, hemos observado que esta droga genera disfunción mitocondrial. Finalmente, mediante un sistema automatizado, hemos puesto a punto un método para evaluar el efecto neurotóxico del cisplatino en el nemátodo y hemos encontrado que la dopamina posee un efecto protector.

The largest U.S. cancer health disparity exists in prostate cancer, with African American men having the highest incidence and mortality rates. The present study evaluated the effects of ZIC2 and the underlying mechanisms in the E006 parental African-American cell line that produces tumors at accelerated growth rates because of the increase of ZIC2 genes in African-American males. We analyzed the experimental research that the overexpression of ZIC2 contributes to progression of prostate cancer. E006AA cells with overexpressed or suppressed ZIC2 were analyzed to determine phenotypic differences, PCR, cell proliferation and immunoblot assays. The expression levels of ZIC2 were analyzed by CRISPR-Cas9, Western blot and proliferation growth curves. We discovered using these experimental techniques to knockout ZIC2, reduced cell proliferation occurred. This research investigated the role of ZIC2 in prostate cancer progression and the effects of the loss or gain of function of ZIC2 by using CRISPR-Cas 9 genome editing technology.

L’aniridie est une pathologie rare principalement due à des mutations hétérozygotes sur PAX6, le gène contrôlant le développement oculaire et le maintien de l’homéostasie cornéenne. Elle est caractérisée par une hypo/aplasie de l’iris, des atteintes de la rétine et du cristallin. La totalité des patients atteints développe aussi une kératopathie associée à l’aniridie (KAA), conduisant à une opacification progressive de leur cornée. La KAA a pour origine un déficit en cellules souches limbiques et diverses perturbations de l’épithélium et du stroma cornéens. Actuellement, il n’existe ni traitement pour soulager efficacement les patients, ni modèle cellulaire pour cette pathologie. Afin de pallier ces manques, le système CRISPR/Cas9 a été utilisé pour intégrer une mutation hétérozygote non-sens dans le gène PAX6 de cellules épithéliales limbiques immortalisées. Les cellules mutées présentent une expression réduite de PAX6 et une modulation de celle de ses gènes cibles, un ralentissement de la prolifération, de la clonogénicité et de la migration ainsi qu’une adhésion accrue. De plus, nous avons montré qu’un traitement de ces cellules avec une protéine recombinante PAX6, portant un peptide de pénétration cellulaire, permet une induction de l’expression endogène de PAX6 et également une restauration partielle du phénotype décrit précédemment. Cette réversion phénotypique valide le modèle d’haploinsuffisance développé et suggère l’implication de PAX6 dans les différentes fonctions restaurées. Les cellules mutées peuvent maintenant être utilisées pour le criblage d’outils thérapeutiques potentiels pour la KAA et pour d’autres défauts liés à une diminution du dosage de PAX6
Aniridia is a rare panocular disease mainly due to PAX6 heterozygous mutations. PAX6 is the master gene of the eye development and it controls also the corneal homeostasis maintenance. Aniridia is characterized by an iris hypo/aplasia, retina and lens defects. All the aniridia patients will also develop an aniridia-related keratopathy (ARK) leading to a progressive corneal opacification. ARK is due to a limbal stem cell deficiency and alterations of corneal epithelium and stroma functions. Unfortunately, there is currently no efficient treatment to relief the patients and no cellular model for this pathology. To remedy these lacks, CRISPR/Cas9 system was used to insert a nonsense mutation into PAX6 gene of immortalized limbal epithelial cells. The mutated cells produce less PAX6 than the wild-type and PAX6 targets gene expression was modulated. They also display a marked slow-down proliferation, clonogenicity, migration and an enhanced adhesion. Moreover, we have shown that addition of recombinant PAX6 protein fused to a cell penetrating peptide to the culture medium was able to activate the endogenous PAX6 expression and to rescue some phenotypic defects of the mutated cells. Therefore, it validates the PAX6 haploinsufficiency model and suggests that PAX6 could be involved in all the rescued functions. The mutated cells can now be used to screen potential therapeutic tools for ARK and for other defects due to low levels of PAX6

Dans ce travail nous avons élaboré une nouvelle méthode de l'analyse de réseau à définir des membres possibles des voies moléculaires qui sont important pour ce phénotype en utilisant la « hit-liste » des expériences « omics » qui travaille dans le réseau intégré (le réseau comprend des interactions protéine-protéine, de transcription, l’acide ribonucléique micro-l’acide ribonucléique messager et celles métaboliques). La méthode tire des sous-réseaux qui sont construit des voies de quatre types les plus courtes (qui ne se composent des interactions protéine-protéine, ayant au minimum une interaction de transcription, ayant au minimum une interaction l’acide ribonucléique micro-l’acide ribonucléique messager, ayant au minimum une interaction métabolique) entre des hit –gènes et des soi-disant « exécuteurs terminaux » - les composants biologiques qui participent à la réalisation du phénotype finale (s’ils sont connus) ou entre les hit-gènes (si « des exécuteurs terminaux » sont inconnus). La méthode calcule la valeur de la centralité de chaque point culminant et de chaque voie dans le sous-réseau comme la quantité des voies les plus courtes trouvées sur la route précédente et passant à travers le point culminant et la voie. L'importance statistique des valeurs de la centralité est estimée en comparaison avec des valeurs de la centralité dans les sous-réseaux construit des voies les plus courtes pour les hit-listes choisi occasionnellement. Il est supposé que les points culminant et les voies avec les valeurs de la centralité statistiquement signifiantes peuvent être examinés comme les membres possibles des voies moléculaires menant à ce phénotype. S’il y a des valeurs expérimentales et la P-valeur pour un grand nombre des points culminant dans le réseau, la méthode fait possible de calculer les valeurs expérimentales pour les voies (comme le moyen des valeurs expérimentales des points culminant sur la route) et les P-valeurs expérimentales (en utilisant la méthode de Fischer et des transpositions multiples).A l'aide de la méthode masterPATH on a analysé les données de la perte de fonction criblage de l’acide ribonucléique micro et l'analyse de transcription de la différenciation terminal musculaire et les données de la perte de fonction criblage du procès de la réparation de l'ADN. On peut trouver le code initial de la méthode si l’on suit le lien https://github.com/daggoo/masterPATH
In this work we developed a new exploratory network analysis method, that works on an integrated network (the network consists of protein-protein, transcriptional, miRNA-mRNA, metabolic interactions) and aims at uncovering potential members of molecular pathways important for a given phenotype using hit list dataset from “omics” experiments. The method extracts subnetwork built from the shortest paths of 4 different types (with only protein-protein interactions, with at least one transcription interaction, with at least one miRNA-mRNA interaction, with at least one metabolic interaction) between hit genes and so called “final implementers” – biological components that are involved in molecular events responsible for final phenotypical realization (if known) or between hit genes (if “final implementers” are not known). The method calculates centrality score for each node and each path in the subnetwork as a number of the shortest paths found in the previous step that pass through the node and the path. Then, the statistical significance of each centrality score is assessed by comparing it with centrality scores in subnetworks built from the shortest paths for randomly sampled hit lists. It is hypothesized that the nodes and the paths with statistically significant centrality score can be considered as putative members of molecular pathways leading to the studied phenotype. In case experimental scores and p-values are available for a large number of nodes in the network, the method can also calculate paths’ experiment-based scores (as an average of the experimental scores of the nodes in the path) and experiment-based p-values (by aggregating p-values of the nodes in the path using Fisher’s combined probability test and permutation approach). The method is illustrated by analyzing the results of miRNA loss-of-function screening and transcriptomic profiling of terminal muscle differentiation and of ‘druggable’ loss-of-function screening of the DNA repair process. The Java source code is available on GitHub page https://github.com/daggoo/masterPATH

Dissertação de mestrado em Biologia Celular e Molecular, apresentada ao Departamento de Ciências da Vida da Faculdade de Ciências e Tecnologia da Universidade de Coimbra
Neural  stem  cells  (NSCs)  reside  in  special  niches  in  the  adult  brain,  including   subventricular  zone  (SVZ)  of  the  lateral  ventricle  and  subgranular  zone  (SGZ)  of  the   dentate  gyrus.  Blood  vessels  are  an  important  coumpound  of  the  neurogenic  niches  as   they  secreate  proteins  such  as  betacellulin  (BTC)  that  stimulate  NSC  proliferation,  self-­‐ renewal  and  differentiation.  BTC  is  a  member  of  the  epidermal  growth  factor  (EGF)  family   of  ligands,  and  has  been  widely  studied  in  many  different  contexts.  Recently,  it  has  been   demonstrated  that,  in  vitro,  BTC  can  induce  NSC  proliferation,  promote  self-­‐renewal  and   prevent  spontaneous  differentiation.  In  vivo,  BTC  can  also  promote  neurogenesis.  BTC  is   released  into  the  neurogenic  niche  by  endothelial  cells  of  the  microvasculature  and  by  the   choroid  plexus  (CP).  It  is  thought  that  BTC  activity  in  NSCs  is  mediated  through  ErbB1  and   ErbB4  receptors  whose  activation  stimulate  the  AKT  and  MEK  signalling  pathways.     In  my  thesis  I  carried  out  analysis  understanding  the  influence  of  BTC  and  other   growth  factors  on  NSCs  in  vitro  using  the  neurosphere  assay  and  measuring  neurospheres   number  and  size.  I  also  studied  the  importance  of  AKT  and  MEK  signalling  pathways  in   NSCs  cultivated  in  medium  supplemented  with  different  growth  factors  including  BTC.  I   observed  an  increase  in  cell  cycle  arrest  when  inhibitors  of  both  signalling  pathways  were   added  together  in  all  culture  conditions.  Studies  to  better  characterize  the  interaction   between  neurogenic  niche  and  BTC  were  also  carried  out  using  newly  developed   visualisation  techniques,  SeeDB  and  CLARITY  that  allowed  us  to  have  a  3D  perspective.  In   parallel,  I  made  BTC  conditional  and  BTC  reporter  contructs  for  generating  mice  using   newly  developed  CRISPR/  Cas  9  technique.  These  mice  will  allow  us  to  better  understand parallel,  I  made  BTC  conditional  and  BTC  reporter  contructs  for  generating  mice  using   newly  developed  CRISPR/  Cas  9  technique.  These  mice  will  allow  us  to  better  understand  the relative importance of BTC in adult NSCs and whether BTC transcription is modulated
As  células  estaminais  neuronais  estão  localizadas  em  nichos  celulares   especializados  no  cérebro  adulto,  incluindo  a  zona  subventricular  (ZSV)  no  ventrículo   lateral  e  a  zona  subgranular  (ZSG)  no  giro  denteado.  Os  vasos  sanguíneos  são  um   componente  importante  dos  nichos  neurogenicos  pois  eles  secretam  proteínas  como  é  o   caso  da  betacelulina  (BTC)  que  estimula  a  proliferação,  auto-­‐renovação,  e  diferenciação   das  células  estaminais  neuronais.  A  BTC  é  um  membro  da  família  de  ligandos  do  factor  de   crescimento  epidermal,  e  tem  sido  amplamente  estudada  em  muitos  contextos  diferentes.   Recentemente,  foi  demonstrado  que,  a  BTC  induz  a  proliferação,  promove  a  auto-­‐ renovação  e  previne  a  diferenciação  espontânea  das  células  estaminais  neuronais,  in  vitro.   In  vivo,  a  BTC  também  promove  a  neurogénese.  A  BTC  é  libertada  no  nicho  neurogénico   pelas  células  endoteliais  da  microvasculatura  e  pelo  plexo  coróide.    Pensa-­‐se  que  a   atividade  da  BTC  nas  células  estaminais  neuronais  é  mediada  pelos  receptores  ErbB1  e   ErbB4,  cuja  ativação  estimula  as  vias  de  sinalização  AKT  e  MEK.   Na  minha  tese,  procedi  a  análises  por  forma  a  perceber  a  influência  da  BTC  e   outros  factores  de  crescimento  nas  células  estaminais  neuronais,  in  vitro,  usando  para  isso   o  ensaio  de  neuroesferas  bem  como  medindo  o  diâmetro  e  contando  o  número  de   neuroesferas.  Também  estudei  a  importância  das  vias  de  sinalização  AKT  e  MEK  nas   células  estaminais  neuronais  postas  em  cultura  em  meio  suplementado  com  diferentes   factores  de  crescimento  incluindo  a  BTC.  Em  todas  as  condições  de  cultura,  eu  observei  um   aumento  da  paragem  do  ciclo  celular  quando  foram  adicionados  os  inibidores  de  ambas  as   vias  de  sinalização  em  conjunto.  Foram  também  levados  a  cabo  estudos  para  melhor   perceber  a  interação  entre  o  nicho  neurogénico  e  a  BTC  usando  para  isso  novas  técnicas  de   visualização  ,  como  é  o  caso  do  SeeDB  e  do  CLARITY  que  nos  permitiu  ter  uma  perspectiva   3D  dessa  mesma  interação.  Em  paralelo,  eu  criei  construções  para  gerar  ratos  condicionais   e  ratos  repórter,  usando  a  nova  técnica  CRISPR/Cas  9.  Estes  ratos  irão  permitir-­‐nos  melhor   perceber  a  importância  relativa  da  BTC  nas  células  estaminais adultas e também perceber onde a transcrição da BTC é modulada

The ability to precisely regulate gene expression patterns and to modify genome sequence in a site-specific manner holds much promise in determining gene function and linking genotype to phenotype. DNA-binding modules have been harnessed to generate customizable and programmable chimeric proteins capable of binding to site-specific DNA sequences and regulating the genome and epigenome. Modular DNA-binding domains from zinc fingers (ZFs) and transcriptional activator-like effectors (TALEs) are amenable to engineering to bind any DNA target sequence of interest. Deciphering the code of TALE repeat binding to DNA has helped to engineer customizable TALE proteins capable of binding to any sequence of interest. Therefore TALE repeats provide a rich resource for bioengineering applications. However, the TALE system is limited by the requirement to re-engineer one or two proteins for each new target sequence. Recently, the clustered regularly interspaced palindromic repeats (CRISPR)/ CRISPR associated 9 (Cas9) has been used as a versatile genome editing tool. This machinery has been also repurposed for targeted transcriptional regulation. Due to the facile engineering, simplicity and precision, the CRISPR/Cas9 system is poised to revolutionize the functional genomics studies across diverse eukaryotic species. In this dissertation I employed transcription activator-like effectors and CRISPR/Cas9 systems for targeted genome regulation and editing and my achievements include: 1) I deciphered and extended the DNA-binding code of Ralstonia TAL effectors providing new opportunities for bioengineering of customizable proteins; 2) I repurposed the CRISPR/Cas9 system for site-specific regulation of genes in plant genome; 3) I harnessed the power of CRISPR/Cas9 gene editing tool to study the function of the serine/arginine-rich (SR) proteins.

Developing targeted genome regulation approaches holds much promise for accelerating trait discovery and development in agricultural biotechnology. Clustered Regularly Interspaced Palindromic Repeats (CRISPRs)/CRISPR associated (Cas) system provides bacteria and archaea with an adaptive molecular immunity mechanism against invading nucleic acids through phages and conjugative plasmids. The type II CRISPR/Cas system has been adapted for genome editing purposes across a variety of cell types and organisms. Recently, the catalytically inactive Cas9 (dCas9) protein combined with guide RNAs (gRNAs) were used as a DNA-targeting platform to modulate the expression patterns in bacterial, yeast and human cells. Here, we employed this DNA-targeting system for targeted transcriptional regulation in planta by developing chimeric dCas9-based activators and repressors. For example, we fused to the C-terminus of dCas9 with the activation domains of EDLL and TAL effectors, respectively, to generate transcriptional activators, and the SRDX repression domain to generate transcriptional repressor. Our data demonstrate that the dCas9:EDLL and dCas9:TAD activators, guided by gRNAs complementary to promoter elements, induce strong transcriptional activation on episomal targets in plant cells. Moreover, our data suggest that the dCas9:SRDX repressor and the dCas9:EDLL and dCas9:TAD activators are capable of markedly repressing or activating, respectively, the transcription of an endogenous genomic target. Our data indicate that the CRISPR/dCas9:TFs DNA targeting system can be used in plants as a functional genomic tool and for biotechnological applications.

This chapter examines the impact of the 2007–08 global financial crisis on the regulation of private investment funds in the United States and in the European Union. It begins with a review of Dodd-Frank, which can be seen as the U.S. movement towards the international consensus that private fund managers should be directly regulated by the national financial regulator. It then considers Dodd-Frank’s repeal of the so-called ‘private adviser exemption’ previously found in the Investment Advisers Act of 1940, along with its exemption of ‘foreign private advisers’ from registration. It also explains the distinction between ‘US advisers’ and ‘non-US advisers’, Dodd-Frank’s compliance requirements for various types of investment advisers, and Rule 204(b)-1, jointly approved by the Securities and Exchange Commission and the Commodity Futures Trading Commission under the Investment Advisers Act. The chapter concludes with an analysis of the Alternative Investment Fund Managers Directive (AIFMD) and future outlook for Dodd-Frank.

This book examines the life of Samuel Wesley, the father of John and Charles Wesley, as a High Church parson in the Church of England. It examines a series of crises in Wesley’s life: his move from Dissent to the Church of England, his abandonment of James II in 1688, his failed ambitions as a parish priest, the imprisonment for debt in 1705, his problematic relations with his bishop and tumultuous marriage to Susanna Wesley, his support for the Tory Convocation measures in 1713 and the haunting of his rectory in Epworth by a poltergeist. Each of these aspects of Wesley’s life showed how awkward his continuing commitment to High Church Toryism was. The book argues that Wesley’s life demonstrates that the Revolution of 1688-9 was not a single event, but a long and protracted experience, reaching, in Wesley’s case, from 1685-1720. The Tory Crisis of Piety of this period was evidence of the Long Glorious Revolution.

Between Realism and Revolt explores urban governance in the “age of austerity”, focusing on the period between the global financial crisis of 2008-9 and the beginning of the global Coronavirus pandemic at the end of 2019. It considers urban governance after the 2008 crisis, from the perspective of governability. How did cities navigate the crisis and the aftermath of austerity, with what political ordering and disordering dynamics at the forefront? To answer these questions it engages with two influential theoretical currents, Urban Regime Theory and Gramscian state theory, with a view to understanding how governance enabled austerity, deflected or intensified localised expressions of crisis, and generated more-or-less successful political alternatives. It develops a comparative analysis of case studies undertaken in the cities of Athens, Baltimore, Barcelona, Greater Dandenong (Melbourne), Leicester, Montreal and Nantes, and concludes by highlighting five characteristics that cut across the cities, unevenly and in different configurations: economic rationalism, weak hegemony, retreat to dominance, weak counter-hegemony and radically contagious politicisations.

Ovid’s poems of exile have found new life not only through Darrieussecq’s translations, but also through the way in which they inform the poetry of Josephine Balmer whose volume The Word for Sorrow includes her ‘transgressions’ of Ovid that take us from the battlefields of the First World War (situated close to the site of Ovid’s exile) to the poet/translator’s present world, as she searches for Ovid in the recesses of the internet and links him to her own family history. On the other side of the Atlantic Averill Curdy, also, is thinking about the reception of Ovid in America, and what his experiences of loss and exile can teach us in the face of tragedies such as 9/11, the financial crisis, and the Iraq war.

For the worlds democracies, the Global Financial Crisis of 2008–9 was catalyst for the most precipitous economic downturn in eight decades. This book examines how the GFC and ensuing Great Recession affected the workings of mass politics in the established democracies. The initial wave of research on the crisis concluded it did little to change the established relationships between voters, parties, and elections. Yet, nearly a decade since the initial shock, we are witnessing a wave of political changes, the extent to which has not been fully explained by existing studies. How did the economic malaise bear on the political preferences of citizens? This book pushes against the received wisdom by advancing a framework for understanding citizen attitudes, preferences, and behaviour. We make two main claims. First, while previous studies of the GFC tend to focus on an immediate impact of the crisis, we argue that economic malaise had a long-lasting impact. In addition to economic shock, we emphasize that economic recovery has a significant impact on citizens assessment of political elites. Second, we argue that unanticipated exogenous shocks like the GFC grant party elites an opening for political manoeuvre through public policy and rhetoric. As a result, political elites have a high degree of agency to shape public perceptions and behaviour. Political parties can strategically moderate citizens economic uncertainty, mobilize/demobilize voters, and alter individuals political preferences. By leveraging data from over 150,000 individuals across over 100 nationally representative post-election surveys from the 1990s to 2017, this book tests these research claims across a range of outcomes, including economic perceptions, policy demands, political participation, and the vote.

Malaysia was once on the cusp of becoming one of the Asian Tigers as a result of the impressively high growth rates recorded in the early 1990s. From 1990 until 1997, the growth rate was above 9 percent per annum on average. This performance came to an end when the economy was struck by the 1997/98 Asian Financial Crisis, the worst economic crisis Malaysia has ever experienced since independence. Things eventually worsened with the onslaught of the 2008/09 Global Financial Crisis, which dragged the Malaysian economy yet into another round of a recession with the growth rate contracting at 1.5 percent in 2009. On hindsight, these two events, which have had a substantial impact on the state of the Malaysian economy, pointed to several urgent calls for economic reforms, such as the need to address structural weaknesses of the economy and to have a growth target which is both sustainable as well as inclusive. When Datuk Seri Najib Razak became the sixth Prime Minister of Malaysia from April 2009 until May 2018, it was clear that a new approach to economic development for Malaysia had to be crafted. Towards this end, he introduced the National Transformation Policy (NTP), so that the economy can be transformed into one that is of high-income and developed status by the year 2020. He also set a new vision for Malaysia, also known as the 2050 National Transformation, or TN50, which is meant to chart a new course for Malaysia to move into the second half of the 21st century. How successful is this transformational agenda? What are the other issues and challenges which need to be addressed? What important lessons can we learn from this transformational journey? This book is an attempt to address these specific questions by assessing Najibs economic plans, policies, programmes and vision which evolved during the nine years of his term as the sixth Prime Minister of Malaysia.

The topic of national development banks was largely neglected in the academic literature for a long period, and was limited to a debate between admirers and detractors of these institutions. Since the 2007/9 financial crisis, interest in and support for these institutions have broadly increased, in developing, emerging, and developed countries alike. The key issues are understanding how such development banks work, what their main aims are, what instruments, incentives, and governance work better in general and in particular contexts, and what are their links with the private financial and corporate sector, as well as with broader government policies. This book aims to provide an in-depth study of several key cases of national development banks (in Brazil, Chile, China, Colombia, Mexico, Germany, and Peru) as well as horizontal issues such as their role in innovation and structural change, infrastructure financing, financial inclusion, environmental sustainability, the countercyclical role of development financing, and the regulatory rules that are best for these institutions. From both a research and a policymaking perspective, this book concludes that development banks can make a significant contribution to development. It analyses their roles, the link with broader economic policies, their governance, and the main instruments they use to perform their functions. The book has important policy implications for countries that have development banks, so they can improve them, but also for countries which do not yet have them, and can learn from best practice should they wish to establish them.

The Black Death of 1348–9 is the most catastrophic event in recorded history, and this study—the Ford Lectures of 2019 at Oxford University—offers a major re-evaluation of its immediate impact and longer-term consequences in England. It draws upon recent inter-disciplinary research into climate and disease; renewed interest among econometricians in the origins of the Little Divergence, whereby economic performance in parts of north-western Europe began to move decisively ahead of the rest of the continent on the pathway to modernity; a close re-reading of case studies of fourteenth-century England; and original new research into manorial and governmental sources. The Black Death is placed within the wider contexts of extreme weather and epidemiological events, the institutional framework of markets and serfdom, and the role of the law in reducing risk and shaping behaviour. The government’s response to the crisis is re-considered to suggest an innovative re-interpretation of the Peasants’ Revolt of 1381. By 1400 the main effects of plague had worked through the economy and society, and their implications for England’s future precocity are analysed. This study rescues the third quarter of the fourteenth century from a little-understood paradox between plague and revolt, and elevates it to a critical period of profound and irreversible change in English and global history.

The volume provides a distinctive new account of British economic life since the Second World War, focusing upon the ways in which successive governments, in seeking to manage the economy, have sought simultaneously to ‘manage the people’: to try and manage popular understanding of economic issues. In doing so, governments have sought not only to shape expectations for electoral purposes but to construct broader narratives about how ‘the economy’ should be understood. The starting point is to ask what goals have been focused upon; how these have been constructed to appeal to the population; and how far the population has accepted these narratives. In its first part, the volume analyses the development of the major narratives from the 1940s onwards. This part covers the notion of ‘austerity’ and its particular meaning in the 1940s; the rise of a narrative of ‘economic decline’ from the late 1950s, and the subsequent attempts to ‘modernize’ the economy; the attempts to ‘roll back the state’ from the 1970s; the impact of ideas of ‘globalization’ in the 1980s and 1990s; and, finally, the way the crisis of 2008/9 onwards was constructed as a problem of ‘debts and deficits’. The second part of the volume then focuses in on four key issues in attempts to ‘manage the people’: productivity, the balance of payments, inflation, and unemployment. It shows how in each case governments have sought to get the populace to understand these issues in a particular light, and have shaped strategies to that end.

Nationalism: Past as Prologue began as a single volume being compiled by Ad Akande, a scholar from South Africa, who proposed it to me as co-author about two years ago. The original idea was to examine how the damaging roots of nationalism have been corroding political systems around the world, and creating dangerous obstacles for necessary international cooperation. Since I (Bruce E. Johansen) has written profusely about climate change (global warming, a.k.a. infrared forcing), I suggested a concerted effort in that direction. This is a worldwide existential threat that affects every living thing on Earth. It often compounds upon itself, so delays in reducing emissions of fossil fuels are shortening the amount of time remaining to eliminate the use of fossil fuels to preserve a livable planet. Nationalism often impedes solutions to this problem (among many others), as nations place their singular needs above the common good. Our initial proposal got around, and abstracts on many subjects arrived. Within a few weeks, we had enough good material for a 100,000-word book. The book then fattened to two moderate volumes and then to four two very hefty tomes. We tried several different titles as good submissions swelled. We also discovered that our best contributors were experts in their fields, which ranged the world. We settled on three stand-alone books:” 1/ nationalism and racial justice. Our first volume grew as the growth of Black Lives Matter following the brutal killing of George Floyd ignited protests over police brutality and other issues during 2020, following the police assassination of Floyd in Minneapolis. It is estimated that more people took part in protests of police brutality during the summer of 2020 than any other series of marches in United States history. This includes upheavals during the 1960s over racial issues and against the war in Southeast Asia (notably Vietnam). We choose a volume on racism because it is one of nationalism’s main motive forces. This volume provides a worldwide array of work on nationalism’s growth in various countries, usually by authors residing in them, or in the United States with ethnic ties to the nation being examined, often recent immigrants to the United States from them. Our roster of contributors comprises a small United Nations of insightful, well-written research and commentary from Indonesia, New Zealand, Australia, China, India, South Africa, France, Portugal, Estonia, Hungary, Russia, Poland, Kazakhstan, Georgia, and the United States. Volume 2 (this one) describes and analyzes nationalism, by country, around the world, except for the United States; and 3/material directly related to President Donald Trump, and the United States. The first volume is under consideration at the Texas A & M University Press. The other two are under contract to Nova Science Publishers (which includes social sciences). These three volumes may be used individually or as a set. Environmental material is taken up in appropriate places in each of the three books. * * * * * What became the United States of America has been strongly nationalist since the English of present-day Massachusetts and Jamestown first hit North America’s eastern shores. The country propelled itself across North America with the self-serving ideology of “manifest destiny” for four centuries before Donald Trump came along. Anyone who believes that a Trumpian affection for deportation of “illegals” is a new thing ought to take a look at immigration and deportation statistics in Adam Goodman’s The Deportation Machine: America’s Long History of Deporting Immigrants (Princeton University Press, 2020). Between 1920 and 2018, the United States deported 56.3 million people, compared with 51.7 million who were granted legal immigration status during the same dates. Nearly nine of ten deportees were Mexican (Nolan, 2020, 83). This kind of nationalism, has become an assassin of democracy as well as an impediment to solving global problems. Paul Krugman wrote in the New York Times (2019:A-25): that “In their 2018 book, How Democracies Die, the political scientists Steven Levitsky and Daniel Ziblatt documented how this process has played out in many countries, from Vladimir Putin’s Russia, to Recep Erdogan’s Turkey, to Viktor Orban’s Hungary. Add to these India’s Narendra Modi, China’s Xi Jinping, and the United States’ Donald Trump, among others. Bit by bit, the guardrails of democracy have been torn down, as institutions meant to serve the public became tools of ruling parties and self-serving ideologies, weaponized to punish and intimidate opposition parties’ opponents. On paper, these countries are still democracies; in practice, they have become one-party regimes….And it’s happening here [the United States] as we speak. If you are not worried about the future of American democracy, you aren’t paying attention” (Krugmam, 2019, A-25). We are reminded continuously that the late Carl Sagan, one of our most insightful scientific public intellectuals, had an interesting theory about highly developed civilizations. Given the number of stars and planets that must exist in the vast reaches of the universe, he said, there must be other highly developed and organized forms of life. Distance may keep us from making physical contact, but Sagan said that another reason we may never be on speaking terms with another intelligent race is (judging from our own example) could be their penchant for destroying themselves in relatively short order after reaching technological complexity. This book’s chapters, introduction, and conclusion examine the worldwide rise of partisan nationalism and the damage it has wrought on the worldwide pursuit of solutions for issues requiring worldwide scope, such scientific co-operation public health and others, mixing analysis of both. We use both historical description and analysis. This analysis concludes with a description of why we must avoid the isolating nature of nationalism that isolates people and encourages separation if we are to deal with issues of world-wide concern, and to maintain a sustainable, survivable Earth, placing the dominant political movement of our time against the Earth’s existential crises. Our contributors, all experts in their fields, each have assumed responsibility for a country, or two if they are related. This work entwines themes of worldwide concern with the political growth of nationalism because leaders with such a worldview are disinclined to co-operate internationally at a time when nations must find ways to solve common problems, such as the climate crisis. Inability to cooperate at this stage may doom everyone, eventually, to an overheated, stormy future plagued by droughts and deluges portending shortages of food and other essential commodities, meanwhile destroying large coastal urban areas because of rising sea levels. Future historians may look back at our time and wonder why as well as how our world succumbed to isolating nationalism at a time when time was so short for cooperative intervention which is crucial for survival of a sustainable earth. Pride in language and culture is salubrious to individuals’ sense of history and identity. Excess nationalism that prevents international co-operation on harmful worldwide maladies is quite another. As Pope Francis has pointed out: For all of our connectivity due to expansion of social media, ability to communicate can breed contempt as well as mutual trust. “For all our hyper-connectivity,” said Francis, “We witnessed a fragmentation that made it more difficult to resolve problems that affect us all” (Horowitz, 2020, A-12). The pope’s encyclical, titled “Brothers All,” also said: “The forces of myopic, extremist, resentful, and aggressive nationalism are on the rise.” The pope’s document also advocates support for migrants, as well as resistance to nationalist and tribal populism. Francis broadened his critique to the role of market capitalism, as well as nationalism has failed the peoples of the world when they need co-operation and solidarity in the face of the world-wide corona virus pandemic. Humankind needs to unite into “a new sense of the human family [Fratelli Tutti, “Brothers All”], that rejects war at all costs” (Pope, 2020, 6-A). Our journey takes us first to Russia, with the able eye and honed expertise of Richard D. Anderson, Jr. who teaches as UCLA and publishes on the subject of his chapter: “Putin, Russian identity, and Russia’s conduct at home and abroad.” Readers should find Dr. Anderson’s analysis fascinating because Vladimir Putin, the singular leader of Russian foreign and domestic policy these days (and perhaps for the rest of his life, given how malleable Russia’s Constitution has become) may be a short man physically, but has high ambitions. One of these involves restoring the old Russian (and Soviet) empire, which would involve re-subjugating a number of nations that broke off as the old order dissolved about 30 years ago. President (shall we say czar?) Putin also has international ambitions, notably by destabilizing the United States, where election meddling has become a specialty. The sight of Putin and U.S. president Donald Trump, two very rich men (Putin $70-$200 billion; Trump $2.5 billion), nuzzling in friendship would probably set Thomas Jefferson and Vladimir Lenin spinning in their graves. The road of history can take some unanticipated twists and turns. Consider Poland, from which we have an expert native analysis in chapter 2, Bartosz Hlebowicz, who is a Polish anthropologist and journalist. His piece is titled “Lawless and Unjust: How to Quickly Make Your Own Country a Puppet State Run by a Group of Hoodlums – the Hopeless Case of Poland (2015–2020).” When I visited Poland to teach and lecture twice between 2006 and 2008, most people seemed to be walking on air induced by freedom to conduct their own affairs to an unusual degree for a state usually squeezed between nationalists in Germany and Russia. What did the Poles then do in a couple of decades? Read Hlebowicz’ chapter and decide. It certainly isn’t soft-bellied liberalism. In Chapter 3, with Bruce E. Johansen, we visit China’s western provinces, the lands of Tibet as well as the Uighurs and other Muslims in the Xinjiang region, who would most assuredly resent being characterized as being possessed by the Chinese of the Han to the east. As a student of Native American history, I had never before thought of the Tibetans and Uighurs as Native peoples struggling against the Independence-minded peoples of a land that is called an adjunct of China on most of our maps. The random act of sitting next to a young woman on an Air India flight out of Hyderabad, bound for New Delhi taught me that the Tibetans had something to share with the Lakota, the Iroquois, and hundreds of other Native American states and nations in North America. Active resistance to Chinese rule lasted into the mid-nineteenth century, and continues today in a subversive manner, even in song, as I learned in 2018 when I acted as a foreign adjudicator on a Ph.D. dissertation by a Tibetan student at the University of Madras (in what is now in a city called Chennai), in southwestern India on resistance in song during Tibet’s recent history. Tibet is one of very few places on Earth where a young dissident can get shot to death for singing a song that troubles China’s Quest for Lebensraum. The situation in Xinjiang region, where close to a million Muslims have been interned in “reeducation” camps surrounded with brick walls and barbed wire. They sing, too. Come with us and hear the music. Back to Europe now, in Chapter 4, to Portugal and Spain, we find a break in the general pattern of nationalism. Portugal has been more progressive governmentally than most. Spain varies from a liberal majority to military coups, a pattern which has been exported to Latin America. A situation such as this can make use of the term “populism” problematic, because general usage in our time usually ties the word into a right-wing connotative straightjacket. “Populism” can be used to describe progressive (left-wing) insurgencies as well. José Pinto, who is native to Portugal and also researches and writes in Spanish as well as English, in “Populism in Portugal and Spain: a Real Neighbourhood?” provides insight into these historical paradoxes. Hungary shares some historical inclinations with Poland (above). Both emerged from Soviet dominance in an air of developing freedom and multicultural diversity after the Berlin Wall fell and the Soviet Union collapsed. Then, gradually at first, right wing-forces began to tighten up, stripping structures supporting popular freedom, from the courts, mass media, and other institutions. In Chapter 5, Bernard Tamas, in “From Youth Movement to Right-Liberal Wing Authoritarianism: The Rise of Fidesz and the Decline of Hungarian Democracy” puts the renewed growth of political and social repression into a context of worldwide nationalism. Tamas, an associate professor of political science at Valdosta State University, has been a postdoctoral fellow at Harvard University and a Fulbright scholar at the Central European University in Budapest, Hungary. His books include From Dissident to Party Politics: The Struggle for Democracy in Post-Communist Hungary (2007). Bear in mind that not everyone shares Orbán’s vision of what will make this nation great, again. On graffiti-covered walls in Budapest, Runes (traditional Hungarian script) has been found that read “Orbán is a motherfucker” (Mikanowski, 2019, 58). Also in Europe, in Chapter 6, Professor Ronan Le Coadic, of the University of Rennes, Rennes, France, in “Is There a Revival of French Nationalism?” Stating this title in the form of a question is quite appropriate because France’s nationalistic shift has built and ebbed several times during the last few decades. For a time after 2000, it came close to assuming the role of a substantial minority, only to ebb after that. In 2017, the candidate of the National Front reached the second round of the French presidential election. This was the second time this nationalist party reached the second round of the presidential election in the history of the Fifth Republic. In 2002, however, Jean-Marie Le Pen had only obtained 17.79% of the votes, while fifteen years later his daughter, Marine Le Pen, almost doubled her father's record, reaching 33.90% of the votes cast. Moreover, in the 2019 European elections, re-named Rassemblement National obtained the largest number of votes of all French political formations and can therefore boast of being "the leading party in France.” The brutality of oppressive nationalism may be expressed in personal relationships, such as child abuse. While Indonesia and Aotearoa [the Maoris’ name for New Zealand] hold very different ranks in the United Nations Human Development Programme assessments, where Indonesia is classified as a medium development country and Aotearoa New Zealand as a very high development country. In Chapter 7, “Domestic Violence Against Women in Indonesia and Aotearoa New Zealand: Making Sense of Differences and Similarities” co-authors, in Chapter 8, Mandy Morgan and Dr. Elli N. Hayati, from New Zealand and Indonesia respectively, found that despite their socio-economic differences, one in three women in each country experience physical or sexual intimate partner violence over their lifetime. In this chapter ther authors aim to deepen understandings of domestic violence through discussion of the socio-economic and demographic characteristics of theit countries to address domestic violence alongside studies of women’s attitudes to gender norms and experiences of intimate partner violence. One of the most surprising and upsetting scholarly journeys that a North American student may take involves Adolf Hitler’s comments on oppression of American Indians and Blacks as he imagined the construction of the Nazi state, a genesis of nationalism that is all but unknown in the United States of America, traced in this volume (Chapter 8) by co-editor Johansen. Beginning in Mein Kampf, during the 1920s, Hitler explicitly used the westward expansion of the United States across North America as a model and justification for Nazi conquest and anticipated colonization by Germans of what the Nazis called the “wild East” – the Slavic nations of Poland, the Baltic states, Ukraine, and Russia, most of which were under control of the Soviet Union. The Volga River (in Russia) was styled by Hitler as the Germans’ Mississippi, and covered wagons were readied for the German “manifest destiny” of imprisoning, eradicating, and replacing peoples the Nazis deemed inferior, all with direct references to events in North America during the previous century. At the same time, with no sense of contradiction, the Nazis partook of a long-standing German romanticism of Native Americans. One of Goebbels’ less propitious schemes was to confer honorary Aryan status on Native American tribes, in the hope that they would rise up against their oppressors. U.S. racial attitudes were “evidence [to the Nazis] that America was evolving in the right direction, despite its specious rhetoric about equality.” Ming Xie, originally from Beijing, in the People’s Republic of China, in Chapter 9, “News Coverage and Public Perceptions of the Social Credit System in China,” writes that The State Council of China in 2014 announced “that a nationwide social credit system would be established” in China. “Under this system, individuals, private companies, social organizations, and governmental agencies are assigned a score which will be calculated based on their trustworthiness and daily actions such as transaction history, professional conduct, obedience to law, corruption, tax evasion, and academic plagiarism.” The “nationalism” in this case is that of the state over the individual. China has 1.4 billion people; this system takes their measure for the purpose of state control. Once fully operational, control will be more subtle. People who are subject to it, through modern technology (most often smart phones) will prompt many people to self-censor. Orwell, modernized, might write: “Your smart phone is watching you.” Ming Xie holds two Ph.Ds, one in Public Administration from University of Nebraska at Omaha and another in Cultural Anthropology from the Chinese Academy of Social Sciences, Beijing, where she also worked for more than 10 years at a national think tank in the same institution. While there she summarized news from non-Chinese sources for senior members of the Chinese Communist Party. Ming is presently an assistant professor at the Department of Political Science and Criminal Justice, West Texas A&M University. In Chapter 10, analyzing native peoples and nationhood, Barbara Alice Mann, Professor of Honours at the University of Toledo, in “Divide, et Impera: The Self-Genocide Game” details ways in which European-American invaders deprive the conquered of their sense of nationhood as part of a subjugation system that amounts to genocide, rubbing out their languages and cultures -- and ultimately forcing the native peoples to assimilate on their own, for survival in a culture that is foreign to them. Mann is one of Native American Studies’ most acute critics of conquests’ contradictions, and an author who retrieves Native history with a powerful sense of voice and purpose, having authored roughly a dozen books and numerous book chapters, among many other works, who has traveled around the world lecturing and publishing on many subjects. Nalanda Roy and S. Mae Pedron in Chapter 11, “Understanding the Face of Humanity: The Rohingya Genocide.” describe one of the largest forced migrations in the history of the human race, the removal of 700,000 to 800,000 Muslims from Buddhist Myanmar to Bangladesh, which itself is already one of the most crowded and impoverished nations on Earth. With about 150 million people packed into an area the size of Nebraska and Iowa (population less than a tenth that of Bangladesh, a country that is losing land steadily to rising sea levels and erosion of the Ganges river delta. The Rohingyas’ refugee camp has been squeezed onto a gigantic, eroding, muddy slope that contains nearly no vegetation. However, Bangladesh is majority Muslim, so while the Rohingya may starve, they won’t be shot to death by marauding armies. Both authors of this exquisite (and excruciating) account teach at Georgia Southern University in Savannah, Georgia, Roy as an associate professor of International Studies and Asian politics, and Pedron as a graduate student; Roy originally hails from very eastern India, close to both Myanmar and Bangladesh, so he has special insight into the context of one of the most brutal genocides of our time, or any other. This is our case describing the problems that nationalism has and will pose for the sustainability of the Earth as our little blue-and-green orb becomes more crowded over time. The old ways, in which national arguments often end in devastating wars, are obsolete, given that the Earth and all the people, plants, and other animals that it sustains are faced with the existential threat of a climate crisis that within two centuries, more or less, will flood large parts of coastal cities, and endanger many species of plants and animals. To survive, we must listen to the Earth, and observe her travails, because they are increasingly our own.

Abstract This chapter discusses the complexity of drought tolerance in wheat focusing the morphological, biochemical, physiological and molecular responses. The breeding approaches, such as traditional and genomics-assisted strategies, for drought tolerance in wheat are described. Future perspectives are also mentioned. Before wheat genome sequencing, it was very difficult to dissect drought tolerance genomic regions because of large genome size and repetitive sequences. But with the availability of sequencing approaches, a large number of genomic resources has become available which extend the scope of utilization of advanced genomics approaches such as GWAM and GS, MutMap+, etc. A new genome editing approach, the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPRassociated protein 9 (Cas9) system, can also be utilized for enhancement of drought tolerance in wheat. Therefore, integration of genomic approaches with precise phenotyping is the need of the hour for improving drought tolerance in wheat.

Background: Last decade over the world, the cardiac disease becomes a leading cause of death. Gene-based therapies become a promising treatment for patients affected by cardiovascular diseases, such as myocardial infarction (MI), arteriosclerosis, heart failure and so on, but also underline the require for reproducible results in preclinical and clinical studies for efficacy and safety. Aim: This book chapter describes the current research prospect of gene therapy for cardiac disease. We focus on the various models to deliver genes using viral, non-viral vector, delivery methods, targets gene, recent clinical trials, inherited cardiomyopathies target genes and Present advances of CRISPR/Cas 9 for cardiovascular gene therapy. We recapitulate some challenges that require being overcome, future directions of gene therapies for cardiac disease. Materials and Methods: All required information regards Lef-7 was generated by exploring the internet search engine like as (PubMed, Wiley, ScienceDirect, CNKI, ACS, Google Scholar, Web of Science, SciFinder, and Baidu Scholar) and libraries. Results: In this book chapter, we focus on the present prospect of gene targets, gene delivery methods, and efficient vector to deliver gene, targets gene, recent clinical trials, inherited cardiomyopathies target genes and present advances of CRISPR/Cas 9 technology for the treatment of cardiac disease using gene therapy. Recent clinical trials require modifying vectors and gene delivery approaches to achieve effective results for cardiac gene therapy. Conclusion: In this book chapter, we integrate a historical perspective with recent advances that will likely affect clinical development in this research area.

Background: Last decade over the world, the cardiac disease becomes a leading cause of death. Gene-based therapies become a promising treatment for patients affected by cardiovascular diseases, such as myocardial infarction (MI), arteriosclerosis, heart failure and so on, but also underline the require for reproducible results in preclinical and clinical studies for efficacy and safety. Aim: This book chapter describes the current research prospect of gene therapy for cardiac disease. We focus on the various models to deliver genes using viral, non-viral vector, delivery methods, targets gene, recent clinical trials, inherited cardiomyopathies target genes and Present advances of CRISPR/Cas 9 for cardiovascular gene therapy. We recapitulate some challenges that require being overcome, future directions of gene therapies for cardiac disease. Materials and Methods: All required information regards Lef-7 was generated by exploring the internet search engine like as (PubMed, Wiley, ScienceDirect, CNKI, ACS, Google Scholar, Web of Science, SciFinder, and Baidu Scholar) and libraries. Results: In this book chapter, we focus on the present prospect of gene targets, gene delivery methods, and efficient vector to deliver gene, targets gene, recent clinical trials, inherited cardiomyopathies target genes and present advances of CRISPR/Cas 9 technology for the treatment of cardiac disease using gene therapy. Recent clinical trials require modifying vectors and gene delivery approaches to achieve effective results for cardiac gene therapy. Conclusion: In this book chapter, we integrate a historical perspective with recent advances that will likely affect clinical development in this research area.

In this study, we assessed the efficiency and off-target activity of the CRISPR/CAS complex with one of the selected guide RNAs using the CIRCLE-seq technology. The gene encoding the human chemokine receptor CCR5 was used as a target sequence for genome editing. The results of this experiment indicate the correct choice of the guide RNA and efficient work of the CRISPR- CAS ribonucleoprotein complex used. CIRCLE-seq technology has shown high sensitivity compared to bioinformatic methods for predicting off-target activity of CRISPR/CAS complexes. We plan to evaluate the efficiency and off-target activity of CRISPR/CAS ribonucleoprotein complexes with other guide RNAs by slightly adjusting the CIRCLE-seq-technology protocol in order to reduce nonspecific DNA breaks and increase the number of reliable reads.

For the effective functioning of supervisory and health monitoring services, it is necessary to introduce modern molecular technologies into their practice. Therefore, the task of developing new effective methods for detecting pathogen, for example HIV, based on CRISPR/CAS genome editing systems, remains urgent. In the present work, guide RNAs and specific oligonucleotides were developed for preliminary amplification of highly conserved regions of the HIV-1 genome. The developed guide RNAs make it possible to detect single copies of HIV-1 proviral DNA in vitro as part of CRISPR/CAS ribonucleoprotein complexes in biological samples after preliminary amplification.

Removal of bacterial endotoxins from solutions of recombinant proteins is one of the most important issues in the preparation of highly purified preparations suitable for in vivo use. An optimal technology for obtaining preparations purified from bacterial endotoxins has been proposed using purification of preparations of recombinant nucleases of the CRISPR/CAS system as an example. Efficacy of developed technology was compared with other available methods. Removal of bacterial endotoxins was carried out using Triton X-114 detergent added to a concentration of 1% to a solution containing the recombinant protein. It was shown that the content of bacterial endotoxins in solutions of purified proteins obtained according to the proposed technology is 0.3–1.5 EU/ml.

The article considers issues related to the crisis of understanding as the key pillar and basis of all other crises. It is proposed to consider the "infology" approach as a primary antidote to obtain a more adequate understanding of the world. This approach can make it possible to educate a new type of person such as homo informationalis, that is an active link in the information world.

Introdução: As plantas são suscetíveis a um grande número de patógenos, incluindo os fungos. Fitopatógenos fúngicos são responsáveis ​​por inúmeras doenças, como ferrugem, oídio, podridão, entre outras. Diferentes estratégias têm sido desenvolvidas para aumentar a resistência fúngica em espécies de plantas com base no conhecimento atual dos mecanismos moleculares envolvidos na interação planta-patógeno. As tecnologias de edição de genoma progrediram rapidamente e se tornaram as ferramentas genéticas mais utilizadas para o melhoramento de plantas. Entre essas, temos a aplicação do sistema formado por repetições palindrômicas curtas, interespaçadas e regularmente agrupadas (CRISPR), e sua proteína associada-9 (Cas9). Objetivo: Apresentar a tecnologia de edição de genoma CRISPR/Cas9 com foco na sua aplicação para o aumento da resistência de plantas á patógenos fúngicos. Metodologia: A pesquisa foi realizada nas bases de dados: PubMed e Scopus. Para alcançar o máximo de precisão na estratégia de busca, utilizou-se os descritores: “plant”, “pathogen”, “fungi or fungus”, “CRISPR”. Resultados: A maioria dos trabalhos envolvendo a resistência de plantas contra patógenos fúngicos estavam relacionados como a capacidade do sistema CRISPR/Cas9 em induzir mutagênese direcionada, com competência em silenciar genes implicados na interação planta-fungo. Foi possível observar vários estudos onde os genes de suscetibilidade da planta hospedeira foram inativados, pois eram necessários para o ciclo de vida do patógeno, demonstrando que a tecnologia é aplicável à resistência a doenças fúngicas em plantas, pois o silenciamento de um determinado gene na planta pode resultar em uma suscetibilidade no fungo. Conclusão: Os resultados demonstram a aplicação vantajosa do sistema CRISPR/Cas9 para o melhoramento de culturas no que diz respeito à resistência a patógenos. O aumento dessa resistência possui um papel importante, pois os fungos fitopatogênicos representam uma ameaça para a produção e o rendimento das safras agrícolas.