Sequencing : Sequencing And Data Generation Essay
Libraries were sequenced on NextSeq 500 platform of Illumina using 2x150 PE Chemistry. The average coverage of the transcriptome on the NextSeq500 platform was ~200x. High quality reads (QV>20) were only used for the RNA-Seq analysis. A total of 11,608,257, 12,184,924, 11,811,964 and 10,592,622 high quality reads were obtained in VS, CONTROL, TR15 and TR30, respectively (Table 2). High quality reads were mapped on to the reference genome using mapping software BWA with optimized parameters as per in our previous study (Sood and Chauhan, 2016).
3.2 Differential gene expression analysis
A total of 55,755 transcripts associated with multiple pathways were expressed in CONTROL, TR15 and TR30 on the basis of FPKM values. 253, 510 and 444 transcripts were expressed uniquely in CONTROL, TR15 and TR30, respectively. 1350 and 1236 transcripts were upregulated and down regulated respectively in TR15 as compared to CONTROL. 3192 and 375 transcripts were upregulated and down regulated respectively in TR15 as compared to TR30. In VS vs CONTROL tissues, about 413 and 419 transcripts were uniquely expressed, whereas 3486 and 191 transcripts were upregulated and down regulated respectively in CONTROL (Fig. 3).
3.3 Pathway analysis
Through KEGG pathway the largest number of transcripts differentially expressed in response to cytokinin treatment were mapped to photosynthesis, Calvin cycle, oxidative phosphorylation, carbohydrate metabolism,…