Transcriptome analysis and gene mining of flowering Chinese cabbage in response to high temperature stress

【Objective】The purpose of the study was to explore the transcriptome analysis and gene mining of flowering Chinese cabbage in response to high temperature stress， and to provide a reference for screening heat-resistant genes of flowering Chinese cabbage in response to high temperature stress and molecular breeding of heat-resistant flowering Chinese cabbage. 【Method】Heat-resistant variety Sijiu-19 flowering Chinese cabbage and heat-sensitive variety Teqingchixin No. 4 flowering Chinese cabbage， two greatly different heat-resistant varieties of flowering Chinese cabbage， were used as experimental materials. After high temperature （42 ℃） stress treatment， transcriptome sequencing （RNASeq） technology was 766 conducted on genes of 24 samples in the shoots and roots of the plants. The differential expression genes（DEGs）were screened， and functional annotation to DEGs was performed using COG， GO， KEGG， KOG， Pfam， Swiss-Prot， eggNOG and Nr biological databases， in order to screen heat shock transcription factors and heat shock proteins in respond to high temperature stress. At the same time， real-time fluorescence quantitative PCR was used to verify important genes of expression and to analyze the cis acting elements of the promoter region. 【Result】A total of 207.18 Gb of clean bases and 47462 DEGs were obtained from 24 samples using RNA-Seq. Only 2663 DEGs were annotated in all 8 functional annotation databases， while the Nr database had the highest number of annotated genes， which were 47324 DEGs， accounting for 99.71% of all genes. Employing eliminating duplicate DEGs from four comparison groups， 15761 DEGs were obtained. The GO functional annotation results showed that DEGs were enriched into three major categories： biological processes， cellular components， and molecular functions. The KEGG signaling pathway enrichment results indicated that the plant hormone signal transduction， carbon metabolism and biosynthesis of amino acids signaling pathways were the most enriched. Under high screening criteria for DEGs with |log₂ Fold Change| ≥8 and P<0.001， a total of 141 DEGs were shared expression in the four comparison groups； 38 genes were specific expression in the Sijiu- 19 flowering Chinese cabbage； and 54 genes were specific expression in the Teqing-chixin No.4 flowering Chinese cabbage. Using databases such as Nr， Pfam and Swiss-Prot， functional annotation analysis was performed on a total of 233 DEGs shared expression and specific expression in two flowering Chinese cabbage varieties， which obtained 9 heat shock transcription factors genes and 25 heat shock proteins genes responsive to high temperature stress. Finally， 10 DEGs highly correlated with response to high temperature stress were selected. The validation results on real-time fluorescence quantitative PCR analysis of 10 DEGs showed that the gene expression trend was consistent with the RNA-Seq data results. The analysis of cis acting elements in the promoter region showed 10 DEGs had multiple cis acting elements that responded to environmental and hormone signals. 【Conclusion】Ten important candidate genes related to response to high temperature induction and significantly up-regulated are screened， and they contain multiple cis acting elements that respond to environmental and hormone signals in the promoter regions.