Genome-wide comparative transcriptome analysis of CMS-D2 and its maintainer and restorer lines in upland cotton

Abstract

Background: Cytoplasmic male sterility (CMS) conferred by the cytoplasm from Gossypium harknessii (D2) is an important system for hybrid seed production in Upland cotton (G. hirsutum). The male sterility of CMS-D2 (i.e., A line) can be restored to fertility by a restorer (i.e., R line) carrying the restorer gene Rf1 transferred from the D2 nuclear genome. However, the molecular mechanisms of CMS-D2 and its restoration are poorly understood.

Results: In this study, a genome-wide comparative transcriptome analysis was performed to identify differentially expressed genes (DEGs) in flower buds among the isogenic fertile R line and sterile A line derived from a backcross population (BC₈F₁) and the recurrent parent, i.e., the maintainer (B line). A total of 1464 DEGs were identified among the three isogenic lines, and the Rf1-carrying Chr_D05 and its homeologous Chr_A05 had more DEGs than other chromosomes. The results of GO and KEGG enrichment analysis showed differences in circadian rhythm between the fertile and sterile lines. Eleven DEGs were selected for validation using qRT-PCR, confirming the accuracy of the RNA-seq results.

Conclusions: Through genome-wide comparative transcriptome analysis, the differential expression profiles of CMS-D2 and its maintainer and restorer lines in Upland cotton were identified. Our results provide an important foundation for further studies into the molecular mechanisms of the interactions between the restorer gene Rf1 and the CMS-D2 cytoplasm.

Keywords: CMS-D2; Circadian rhythm; RNA-seq; Restorer gene; Upland cotton.

Fig. 1

Gene ontology classification (a) and COG functional categories (b) of unigenes

Fig. 2

Distribution of the differentially expressed genes on different chromosomes. a Location distribution of DEGs on different chromosomes. b DEG numbers on different chromosomes. The Y-axis represents different chromosomes. xis and numbers behind each bar represent the DEG numbers on each chromosome

Fig. 3

Venn diagram showing the distribution of unique and common DEGs among three comparisons

Fig. 4

Heatmap showing the FPKM values of DEGs on Chr_D05 and DEGs related to circadian rhythm. The FPKM values for the DEGs in the three samples were used for hierarchical analysis. The heatmap shows the expression abundance of the DEGs. The colors correspond to FPKM values, ranging from blue (low expression) to red (high expression). Those genes in green boxes represent DEGs related to circadian rhythm

Fig. 5

qRT-PCR analysis of gene expression compared with the RNA-seq data. The gray columns represented the relative expression levels of the genes; the dotted lines represent the RNA-seq reads. A: sterile line, B: maintainer line, R: restorer line