Molecular landscape of cotton fibers in early elongation



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Cotton fibers are the dominant source of natural fibers used in the textile industry and contribute significantly to the world economy. Adverse environmental conditions negatively affect fiber characteristics, especially when the fibers are in the elongation phase of development. Improvement in the yield and quality of cotton fibers requires the identification of the molecular networks involved in fiber development. In this research, we have analyzed cotton fiber transcriptome and proteome in the early elongation phase using RNA-Seq and mass spectrometry, to identify the genes and proteins involved in different pathways altered during fiber development. TruSeq RNA sample preparation kit was used to prepare cDNA libraries from RNA extracted from Upland cotton cultivar TM-1 cotton fibers samples at 3 and 5 days post-anthesis (DPA). These libraries were sequenced using 151 bp, paired-end Illumina sequencing which produced 6,570,054 and 7,063,378 reads from 3 DPA and 5 DPA libraries, respectively. The de novo assembly of these raw reads with NGen identified 20,270 contigs in 3 DPA and 20,339 contigs in 5 DPA. The raw reads were mapped on the reference early fiber transcriptome and differential gene expression was estimated by QSeq (DNASTAR genome suite). A total of 3,177 transcripts were recognized as differentially expressed with 95% probability and at least a 2-fold change between 3 DPA and 5 DPA. These transcripts were annotated using Mercator and mapped to biological pathways using MapMan version 3.5.1. The up-regulated transcripts at 5 DPA belonged to cell-wall modification, phospholipid and sphingolipid synthesis, solute and water transporters, cytoskeletal elements and phytohormone categories; processes that were involved in cell wall extension, hinting that the fibers at this stage are involved in loosening the cell wall in anticipation of the rapid fiber elongation beyond 5 DPA. The NGen assembled transcriptomes from 6 stages of fiber development, 3 DPA, 5 DPA, 11 DPA, 17 DPA, 21 DPA and 24 DPA, were re-assembled using CAP3 assembly software to generate the cotton fiber transcriptome. We have also created a reference fiber proteome database from the fiber specific transcriptome containing 374,562 possible protein sequences for protein identification. This is the first report on the generation of a transcriptome and a proteome of the elongating cotton fibers using RNA-Seq. These databases form a significant source of information, and will contribute towards research on the improvement of cotton fiber characteristics.

This thesis won 2nd Place in the Texas Tech University Outstanding Thesis and Dissertation Award, Biological Life Sciences, 2013.



Cotton fibers, Transcriptomics, RNA-seq, Mass spectrometry