Project description:The total RNA were extracted from pooled tissues of leaves and flowers from several plants of chickpea (Cicer arietinum) using TRIzol reagent (Invitrogen) according to the manufacturer's instructions. Then small RNAs ranging in 18–30 nucleotides were size fractionated electrophoretically, isolated from the gel, ligated with the 5′ and 3′ RNA adapters. The ligated product was reverse transcribed and subsequently amplified using 10–12 PCR cycles. The purified PCR product was sequenced using Illumina Genome Analyzer II. The qualified reads were used to predict microRNAs and phased small interfering RNAs from chickpea. Identification of microRNAs and phased small inferfering RNAs in chickpea (Cicer arietinum) by analyzing small RNA sequencing profiles of leaves and flowers using Illumina GAII.
Project description:The total RNA were extracted from pooled tissues of leaves and flowers from several plants of chickpea (Cicer arietinum) using TRIzol reagent (Invitrogen) according to the manufacturer's instructions. Then small RNAs ranging in 18–30 nucleotides were size fractionated electrophoretically, isolated from the gel, ligated with the 5′ and 3′ RNA adapters. The ligated product was reverse transcribed and subsequently amplified using 10–12 PCR cycles. The purified PCR product was sequenced using Illumina Genome Analyzer II. The qualified reads were used to predict microRNAs and phased small interfering RNAs from chickpea.
Project description:Size fractionated small RNA from total RNA extracts of Cicer arietinum leaves and from Nicotiana benthamiana infected by Cymbidium ringspot virus were mixed in a ratio of 1000 to 1 in amount, respectively. The RNA was ligated to adapters, purified again and reverse transcribed. After PCR amplification the sample was subjected to Illumina high throughput pyrosequencing. The kit used is TrueSeq Small RNA kit Please see www.illumina.com for details of the sequencing technology. Short RNA fractionation and characterization
Project description:Chickpea (Cicer arietinum L.) seeds are valued for their nutritional scores and limited information on the molecular mechanisms of chickpea fertilization and seed development is available. In the current work, comparative transcriptome analysis was performed on two different stages of chickpea ovules (pre- and post-fertilization) to identify key regulatory transcripts. Two-staged transcriptome sequencing was generated and over 208 million reads were mapped to quantify transcript abundance during fertilization events. Mapping to the reference genome showed that the majority (92.88%) of high-quality illumina reads were aligned to the chickpea genome. Reference-guided genome and transcriptome assembly yielded a total of 28,783 genes. Of these, 3399 genes were differentially expressed after the fertilization event. These involve up-regulated genes including LOC101500970, LOC101506539 and down-regulated genes LOC101493897, LOC101491695 and so on. Transcription factor families including UDP-glucuronyltransferase, NAC transcription factor, heat shock transcription factor, and auxin-responsive transcription factor were also found to be activated after fertilization. Activation of these genes and transcription factors results in the accumulation of carbohydrates and proteins by enhancing their trafficking and biosynthesis. Total 17 differentially expressed genes, were randomly selected for qRT-PCR for validation of transcriptome analysis and showed statistically significant correlations with the transcriptome data. Our findings provide insights into the regulatory mechanisms underlying changes in fertilized chickpea ovules. This work may come closer to a comprehensive understanding of the mechanisms that initiate developmental events in chickpea seeds.
Project description:Size fractionated small RNA from total RNA extracts of Cicer arietinum leaves and from Nicotiana benthamiana infected by Cymbidium ringspot virus were mixed in a ratio of 1000 to 1 in amount, respectively. The RNA was ligated to adapters, purified again and reverse transcribed. After PCR amplification the sample was subjected to Illumina high throughput pyrosequencing. The kit used is TrueSeq Small RNA kit Please see www.illumina.com for details of the sequencing technology.