云南栘[木衣]MADS-box基因家族鉴定与表达分析

Identification and expression analysis of MADS-box gene family in Docynia delavayi (Franch.) Schneid.

摘要:

MADS-box基因家族是一类重要的转录因子家族，在调控植物的生长发育、信号传导等过程中发挥着重要作用。为研究云南栘[木衣][Docynia delavayi（Franch.） Schneid.]MADS-box基因家族的特征及其在种子不同萌发时期的表达情况，本研究以云南栘[木衣]不同萌发时期的种苗为材料，在转录组测序的基础上利用生物信息学方法从云南栘[木衣]转录组数据库中筛选MADS-box转录因子，分析其理化性质、蛋白保守基序、系统进化及表达模式，并采用实时荧光定量PCR （quantitative real-time polymerase chain reaction，qRT-PCR）实验验证云南栘[木衣]MADS-box基因家族成员在种子不同萌发时期的表达情况。在云南栘[木衣]转录组数据中共鉴定出81个MADS-box转录因子，其编码的氨基酸序列分子量分布范围在6 211.34–173 512.77 Da之间，等电点介于5.21—10.97之间。系统进化分析显示，81个云南栘[木衣]MADS-box基因可分为15个亚组，其中DdMADS27、DdMADS42、DdMADS45、DdMADS46、DdMADS53、DdMADS61、DdMADS76、DdMADS77和DdMADS79可能参与对云南栘[木衣]胚珠的发育调控。结合云南栘[木衣]种子转录组数据与qRT-PCR实验分析发现，DdMADS25和DdMADS42可能参与调控种子发育，DdMADS37和DdMADS38可能对种子休眠有负调控作用。前人报道中MIKC^*亚组多参与调控花器官发育，本研究首次发现MIKC^*亚组的转录因子在种子萌发前期具有较高表达量，由此推测MIKC^*亚组在种子萌发过程中起到调控作用。为验证该推测准确性，挑选了MIKC^*亚组的DdMADS60和DdMADS75进行qRT-PCR实验，实验结果与转录组测序的表达趋势一致。本研究可为进一步从分子进化角度研究云南栘[木衣]MADS-box基因家族的生物学功能提供参考。

Abstract:

MADS-box gene family is a significant transcription factor family that plays a crucial role in regulating plant growth, development, signal transduction, and other processes. In order to study the characteristics of MADS-box gene family in Docynia delavayi (Franch.) Schneid. and its expression during different stages of seed germination, this study used seedlings at different stages of germination as materials and screened MADS-box transcription factors from the transcriptome database of D. delavayi using bioinformatics methods based on transcriptome sequencing. The physical and chemical properties, protein conservative motifs, phylogenetic evolution, and expression patterns of the MADS-box transcription factors were analyzed. Quantitative real-time PCR (qRT-PCR) was used to verify the expression of MADS-box gene family members during different stages of seed germination in D. delavayi. The results showed that 81 genes of MADS-box gene family were identified from the transcriptome data of D. delavayi, with the molecular weight distribution ranged of 6 211.34-173 512.77 Da and the theoretical isoelectric point ranged from 5.21 to 10.97. Phylogenetic analysis showed that the 81 genes could be divided into 15 subgroups, among which DdMADS27, DdMADS42, DdMADS45, DdMADS46, DdMADS53, DdMADS61, DdMADS76, DdMADS77 and DdMADS79 might be involved in the regulation of ovule development in D. delavayi. The combination of the transcriptome data and the qRT-PCR analysis results of D. delavayi seeds indicated that DdMADS25 and DdMADS42 might be involved in the regulation of seed development, and that DdMADS37 and DdMADS38 might have negative regulation effects on seed dormancy. Previous studies have reported that the MIKC^* subgroup is mainly involved in regulating flower organ development. For the first time, we found that the transcription factors of the MIKC^* subgroup exhibited a high expression level at the early stage of seed germination, so we speculated that the MIKC^* subgroup played a regulatory role in the process of seed germination. To verify the accuracy of this speculation, we selected DdMADS60 and DdMADS75 from the MIKC^* subgroup for qRT-PCR experiments, and the experimental results were consistent with the expression trend of transcriptome sequencing. This study provides a reference for further research on the biological function of D.delavayi MADS-box gene family from the perspective of molecular evolution.