Academic literature on the topic 'Germplasm resources'

An in vitro method for the establishment and storage of over 200 Saccharum spp. hybrid clones was developed that involved only 1 medium for shoot development and multiplication, without decontamination procedures. Apical buds from the axils of developing leaves surrounding the apical meristem were cultured on medium containing the plant growth regulators benzylamino purine and kinetin which regenerated multiple shoots. Shoots transferred to a medium containing naphthalene acetic acid and 60 g L"1 sucrose, developed roots. In vitro plants were transferred to a half strength Murashige and Skoog medium without plant growth regulators and placed in storage at 18°C. After 12 months of storage plants were transferred to a fresh medium and returned to storage. The genetic integrity of clones based on phenotypic assessment and isozyme patterns was not affected by in vitro culture after 3, 6 and 12 months storage at l 8°C.

Ctenocephalides felis is a major pest of companion animals worldwide. This project aimed to generate novel genetic resources for C. felis and develop tools to aid drug-target identification and validation. Sample handling methods were assessed and candidate reference genes validated, to ensure quality of RNA samples and reliable gene expression normalisation. Piercing C. felis samples prior to storage in RNAlater ensured RNA integrity was maintained over time. Glyceraldehyde 3-phosphate dehydrogenase , 60S ribosomal protein L19 and elongation factor-1α were demonstrated as stable reference genes across all comparisons tested. A C. felis transcriptome encompassing multiple developmental stages, sexes and tissues was sequenced and de novo assemblies produced with two assemblers, Trinity and Oases. Each assembly contained >100000 contigs. Annotation of the assemblies generated functional insight, such as top BLAST hits, GO annotations and signal peptide predictions. The Trinity assembly was deemed the highest quality and searched for genes of interest, involved in development. Expression analysis of selected transcripts across stadia gave insight into developmental processes, and demonstrated the utility of the transcriptome. This study was the first to demonstrate that C. felis can mount an RNAi response upon exposure to dsRNA. Knockdown of glutathione S-transferase σ (GSTσ), was demonstrated in adult C. felis: ≈80 % knockdown following microinjection of dsGSTσ; ≈64 % knockdown after soaking in dsGSTσ; ≈96 % knockdown after continuous feeding on dsGSTσ. RNAi machinery was identified in C. felis. siRNAi pathway components, Dicer 2 and Argonaute 2, were upregulated following dsRNA exposure. Dicer 2 was knocked-down by soaking in dsDicer2, although results of an “RNAi of RNAi” experiment were inconclusive. Transcripts encoding machinery putatively involved in dsRNA uptake and breakdown were also identified. Through these studies, this project has generated novel insights into C. felis biology and opened up new avenues for research.