Project Summaries

07-161  Project Manager: D. C. Jones


Z. Jeffrey Chen, University of Texas

For 2012 we performed mapping, quantification of expression, statistical analysis of differential expression of RNAseq reads from 5 stages (10, 15, 18, 21, and
28 DPAs) of two allotetraploid species, G. hirsutum (Gh) and G. barbadense (Gb) using our A and D EST references. Comparison between stages showed similar patterns in both Gh and Gb, while greater differences in gene expression in early stages (10 and 15 DPAs) exist between species.

We developed and modified previous A and D EST references in the following way: Since D genome has been sequenced and its transcripts are available, we adopted D transcripts from JGI, which totals 77,267. Then we performed reciprocal blast D transcripts between A EST contigs that we assembled and screened for insignificant matches. We considered the insignificant matches as genome-specific expression: the number of A-specific EST contigs which could not be found in D transcriptome is 34,305 and that of D specific transcripts that are not found in our A EST collection is 9,202. Therefore, the reference is composed of 3 parts: A and D-shared transcripts (68,065, 61%), A-specific transcripts (34,305, 30.7%), and D-specific transcripts (9,202, 8.3%), making a total of 111,572 transcripts.

Differential gene expression between samples was performed using the samWrapper package at R for 3 biological replicates followed by multiple testing. We performed the test for five stages in two species (Gh and Gb) and for two species in five stages.  In Gh, the comparison between 10 and 15 DPA showed 11,709 genes of up-regulation in 10 DPA and 6,218 genes of up-regulation in 15 DPA. Between 15 and 18 DPA, 5,863 genes were up-regulated in 15 DPA, while 3,887 genes were up-regulated in 18 DPA. Between 18 and 21 DPA, 3 genes were up-regulated in 18 DPA, while 29 genes were up-regulated in 21 DPA.
Between 21 and 28 DPA, 24,900 genes were up-regulated in 21 DPA, while 7,158 genes were 2 up-regulated in 28 DPA. Between 28 and 10 DPA, 8,056 genes were up-regulated in 28 DPA, while 26,711 genes were up-regulated in 10 DPA. Thus, the comparison between 18 and 21 DPA showed most similar gene expression, while comparison between 28 and 10 DPA showed most different gene expression. Similar patterns were found in Gb, but there were differences in gene expression between Gb and Gh at elongation stages.

We re-estimated the GNPs between JGI D transcripts and our A reads. We mapped 454 raw reads from A transcriptome to JGI D transcripts using runMapping function at Newbler. With greater than 4X coverage and less than 10% variation, we selected high quality GNPs. A total of 137,923 GNPs were identified in 12,712 genes between D and A transcripts. These genes include epigenetic regulators, transcription factors, and hormone-related genes. This GNP table will be used for quantification of allelic expression.


Project Year: 2012

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