谷子SiCBL3对非生物胁迫响应特征分析

doi:10.13304/j.nykjdb.2022.0566

摘要/Abstract

摘要：

钙调磷酸酶Ｂ蛋白（calcineurin B-like proteins， CBL）在植物逆境应答中起重要作用。前期研究表明谷子SiCBL3受干旱和盐的强烈诱导，因此对其进行系统分析。检索发现SiCBL3位于3号染色体，有2个转录本，均编码226个氨基酸，含3个典型的EF-hand功能域。亚细胞定位显示，SiCBL3蛋白主要位于液泡膜上。荧光实时定量PCR分析表明，SiCBL3广泛参与谷子苗期对PEG、盐、高温、低温和ABA等多种非生物逆境胁迫响应；SiCBL3在正常抽穗和灌浆期表达量较高，且在谷子拔节、抽穗和灌浆期干旱胁迫下被大量诱导。组织表达分析揭示，SiCBL3主要在地上部的倒2叶、茎杆、叶鞘、穗等器官中表达，而在地下部根中表达量较低；不同生育阶段SiCBL3在茎叶中表达量较高，在根中较低；灌浆期SiCBL3主要在倒2叶、茎、穗轴、叶鞘、籽粒等地上部组织中表达，而且在干旱胁迫下被大量诱导。这些结果表明，SiCBL3主要在地上部组织中高效表达，在生长中后期积极地参与植物对干旱等逆境胁迫响应，特别是在灌浆期干旱胁迫应答中起重要作用。研究结果为解析谷子CBL家族基因在逆境应答中的功能奠定基础。

关键词: 谷子, CBL, 非生物逆境, 表达分析

Abstract:

Calcineurin B-like protein （CBL） plays an important role in plant stress response. Previous studies showed that SiCBL3 was strongly induced by drought and salt， so this study further analyzed its function. Retrieval revealed that SiCBL3 was located on chromosome 3， had 2 transcripts， encoded 226 amino acids， and contained 3 typical EF-hand domains. Subcellular localization speculated that SiCBL3 protein was mainly located on the vacuolar membrane. Real-time PCR analysis showed that SiCBL3 responsed to PEG， salt， high temperature， low temperature and ABA in the foxtail millet seedling stage. The analysis of drought stress at different growth stages showed that SiCBL3 was highly expressed at heading and filling stage under normal conditions， and was induced by drought at jointing， heading and filling stage. Tissue expression analysis revealed that SiCBL3 was mainly expressed in the top 2^nd leaves， stems， leaf sheaths， panicles and other aboveground organs， while the expression level in roots was low. The analysis of different growth stages also showed that the expression of SiCBL3 was higher in stems and leaves， but lower in roots. Subsequent tissue analysis at the filling stage also showed that SiCBL3 was mainly expressed in aboveground tissues such as top 2^nd leaf， stem， spike-stalk， leaf sheath and grain， and was strongly induced under drought stress. These results revealed that SiCBL3 was highly expressed in the aboveground tissues， and participated in the response to drought and other stresses in the middle and late stages， especially played an important role in the response to drought stress at the filling stage. This study laid a foundation for further function analysis of CBL family genes in foxtail millet.

Key words: foxtail millet, CBL, abiotic stress, expression analysis

中图分类号:

S515

赵晋锋, 余爱丽, 李颜方, 杜艳伟, 王高鸿, 王振华. 谷子SiCBL3对非生物胁迫响应特征分析[J]. 中国农业科技导报, 2022, 24(11): 68-75.

Jinfeng ZHAO, Aili YU, Yanfang LI, Yanwei DU, Gaohong WANG, Zhenhua WANG. Response Characteristics of SiCBL3 to Abiotic Stresses in Foxtail Millet[J]. Journal of Agricultural Science and Technology, 2022, 24(11): 68-75.

图/表 7

表1 本文实验使用引物

Table 1 Primers used in this study

引物 Primer	上游引物 Forward primer （5’-3’）	下游引物 Reverse primer （5’-3’）
A	TGTCTAGACTCGAGGGATCCATGGTGCAGTGCCTG-GAC	GGCCGCTGTACACATGGATCCGTTGTCATCAACTTGAGAAT-GAAAG
B	GACGCACAATCCCACTATCC	GGGTGAGCTTGCCGTAGGTG
SiCBL3	AGCCTCCCAGATTTGAGACAG	AAGCATTCATCACTGACGGC
β-actin	CAGTGGACGCACAACAGGTAT	AGCAAGGTCAAGACGGAGAAT

表 2 SiCBL3基因启动子区域顺式元件预测

Table 2 Putative cis-elements in the promoter of SiCBL3

顺式元件Cis-element	预测元件Predicted cis-element
植物激素应答 Auxin-responsive	脱落酸（ABRE）、茉莉酮酸甲酯（CGTCA-motif、TGACG-motif）、赤霉素（GARE-motif）、水杨酸（TCA-element） Abscisic acid （ABRE），methyl jasmonate （CGTCA- motif， TGACG-motif），gibberellin （ GARE-motif）， salicylic acid （TCA-element）
逆境应答 Stress-responsive	干旱诱导MBS结合位点、低温响应（LTR） MBS binding sites induced by drought， response to low temperature （LTR）
光应答 Light-responsive	G-box， Box 4， GT1-motif， TCCC-motif
其他应答 Others responsive	昼夜节律、玉米蛋白代谢调控（O₂-site）、厌氧诱导（ ARE） Circadian， metabolic regulationof corn protein （O₂-site）， anaerobic induction （ARE）

图 1 SiCBL3亚细胞定位

Fig. 1 Sub-cellular localization of SiCBL3

图 2 SiCBL3苗期不同逆境胁迫表达分析注：*和**分别表示与处理0 h相比在 P<0.05和P<0.01水平差异显著。

Fig. 2 Expression analysis of SiCBL3 under different stresses in seedlingNote：* and ** indicate significant difference compared with 0 h treatment at P<0.05 and P<0.01 levels， respectively.

图 3 SiCBL3 组织表达分析注：*和**分别表示与旗叶间差异在 P<0.05和P<0.01水平差异显著。

Fig. 3 Expression analysis of SiCBL3 in different tissuesNoe： * and ** indicate significant difference compared with flag leaf at P<0.05 and P<0.01 levels， respectively.

图 4 关键生育阶段干旱胁迫下SiCBL3表达分析注：*和**分别表示与茎叶相比在 P<0.05和P<0.01水平差异显著。

Fig. 4 Expression analysis of SiCBL3 under drought stress at key growth stagesNote： * and ** indicate significant difference compared with stem-leaf at P<0.05 and P<0.01 levels， respectively.

图 5 灌浆期旱胁迫下SiCBL3组织表达分析注：*和**分别表示与对照相比在 P<0.05和P<0.01水平差异显著。

Fig. 5 Expression analysis of SiCBL3 in different tissues under drought stress at filling stageNote： * and ** indicate significant difference compared with CK at P<0.05 and P<0.01 levels， respectively.

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