Project description:Total RNA was isolated from different tissues (leaf, stem and flesh, rind and placenta of the fruits) using TRIzol reagent and small RNA libraries were generated from four cucurbit species: bottle gourd (Lagenaria siceraria (accession Grif 1617 collection from India)), Cucurbita moschata (accession Grif 14244 Early Butternut) Cucurbita pepo (accession NSL98075 Table King), and watermelon (Citrullus lanatus var. lanatus) (PI 438676 Charleston Grey) by pooling equimolar amounts of total RNA from the aforesaid tissues. Construction of small RNA libraries from size fractionated RNA was carried out as described previously. In brief, small RNA fractions of 18–28 nt were isolated from 15% denaturing polyacrylamide gels and sequentially ligated to 5′ and 3′ RNA adapters. Small RNAs ligated with adapters were converted to DNA by RT-PCR following Solexa protocol. The final PCR product was gel purified and sequenced by Genome Analyser II (Illumina).
Project description:Total RNA was isolated from different tissues (leaf, stem and flesh, rind and placenta of the fruits) using TRIzol reagent and small RNA libraries were generated from four cucurbit species: bottle gourd (Lagenaria siceraria (accession Grif 1617 collection from India)), Cucurbita moschata (accession Grif 14244 Early Butternut) Cucurbita pepo (accession NSL98075 Table King), and watermelon (Citrullus lanatus var. lanatus) (PI 438676 Charleston Grey) by pooling equimolar amounts of total RNA from the aforesaid tissues. Construction of small RNA libraries from size fractionated RNA was carried out as described previously. In brief, small RNA fractions of 18–28 nt were isolated from 15% denaturing polyacrylamide gels and sequentially ligated to 5? and 3? RNA adapters. Small RNAs ligated with adapters were converted to DNA by RT-PCR following Solexa protocol. The final PCR product was gel purified and sequenced by Genome Analyser II (Illumina). Examination of small RNA transcriptomes in four plants species using Illumina/Solexa GA-II.
Project description:Cucurbita pepo is high susceptible to Zucchini yellow mosaic virus (ZYMV) and the resistance found in several wild species does not provide complete or broad-spectrum resistance. In this study, a source of tolerance introgressed in C. pepo (381e) from C. moschata, in True French (TF) background, was investigated 12 days after inoculation (DPI) at transcriptomic and genomic levels. A comparative RNA-seq experiment, allowed to evaluate more than 33,000 expressed transcripts and to identify 146 differentially expressed genes (DEGs) in 381e, mainly involved in photosynthesis, transcription, cytoskeleton organization and callose synthesis. By contrast, the susceptible line True French triggered oxidative processes related to response to biotic stimulus and two synaptotagmin (SYTA) genes, key regulators of plant virus intercellular movement. Finally, transcripts mapping allowed the identification of two regions rich in SNPs (Single Nucleotide Polymorphisms) on linkage group 1 and linkage group 8, putatively introgressed from C. moschata, containing a putative disease resistance protein (CNL gene) exclusively expressed in 381e. In conclusion, 381e transcriptome analysis confirmed a globally improved plant fitness by reducing the effect of viral infection and showed the activation of genes putatively involved in tolerance to ZYMV. Our work provides new insight into plant virus recovery process to ZYMV.