羊草种子萌发相关基因的筛选及表达分析

doi:10.11686/cyxb2017502

摘要/Abstract

摘要： 前期实验表明,变温处理(28 ℃ 12 h/16 ℃ 12 h)可显著提高羊草种子的萌发率,且羊草种子萌发中的第1天是接受变温信号的关键时期。以此为研究基础,结合羊草种子变温萌发的转录组测序数据,针对羊草种子萌发初期对变温处理的响应筛选出与种子萌发、休眠及低温相关的基因24个,利用测序结果中这些基因的RPKM值制作基因表达热图并分析其表达差异。以萌发率高、低的两种羊草种质的种子为材料,对24个基因在恒温12 h(28 ℃)和变温1 d(28 ℃ 12 h/16 ℃ 12 h)萌发处理中的表达分别进行了定量分析。结果表明,与恒温对照相比,变温处理12 h后,表达明显上调的基因有SAIN1,PP2C62,EXPB3,EXPB4,GA3ox,EXPA2和EXPA7,而表达明显下调的基因有bHLH49,GID1,ABI8,Chi1,11833,CBF3,NAC2,PP2C72,SAIN2和5423。通过进一步分析相关基因在高、低萌发率两个种质中表达的差异,筛选出其中可能与羊草种子萌发相关的基因有几丁质酶基因Chi1,转录因子基因CBF3,羊草新基因5423,赤霉素合成基因GA3ox,细胞松弛素蛋白基因EXPB4和羊草新基因SAIN1,将为下一步阐明羊草种子萌发的分子作用机理奠定基础。

关键词: 羊草, 种子萌发, 变温, 基因表达

Abstract: Previous studies have suggested that the variable temperature (28 ℃ 12 h/16 ℃ 12 h) can significantly increase the germination rate of Leymus chinensis seed, and that the first day of germination was key for responses to variable temperature. A transcriptome analysis of L. chinensis seed germination under variable temperature treatment was performed. Based on sequencing, 24 genes related to seed germination, low temperature and dormancy were selected for heat map analysis using the reads per kilo base per million mapped reads (RPKM) value. In addition, under the treatment of constant temperature (28 ℃) for 12 h and variable temperature (28 ℃ 12 h/16 ℃ 12 h) for 1 d, the relative expression of the selected genes in two L. chinensis genotypes (high and low germination rate) were quantified by qRT-PCR. Our results indicated that the expression of SAIN1, PP2C62, EXPB3, EXPB4, GA3ox, EXPA2 and EXPA7 were significantly up-regulated in the seeds treated at 16 ℃ for 12 h, while the expression of bHLH49, GID1, ABI8, Chi1, 11833, CBF3, NAC2, PP2C72, SAIN2 and 5423 were significantly down-regulated, compared with the constant temperature (28 ℃) control. Further analysis of the gene expression in the two genotypes suggested that Chi1, CBF3, GA3ox, EXPB4, 5423 and SAIN1 were the putative genes related to germination in L. chinensis.This research may lay the foundation for further studies on the molecular mechanisms in seed germination of L. chinensis.

Key words: Leymus chinensis, seed germination, variable temperature treatment, gene expression

刘书, 刘盼盼, 杨伟光, 齐冬梅, 李晓霞, 刘公社. 羊草种子萌发相关基因的筛选及表达分析[J]. 草业学报, 2018, 27(11): 58-66.

LIU Shu, LIU Pan-pan, YANG Wei-guang, QI Dong-mei, LI Xiao-xia, LIU Gong-she. The identification and expression analysis of genes related to seed germination in Leymus chinensis[J]. Acta Prataculturae Sinica, 2018, 27(11): 58-66.

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