小麦GzCIPK7-5B基因的生物信息学及表达分析

doi:10.16035/j.issn.1001-7283.2023.04.012

摘要/Abstract

摘要：

CIPK是植物钙感受器钙调磷酸酶B类似蛋白特定靶向的一类丝氨酸/苏氨酸蛋白激酶，在钙离子信号转运中起到重要作用。为探究GzCIPK7-5B基因在小麦中的功能，利用RT-PCR方法从小麦品种贵紫麦1号中克隆得到GzCIPK7-5B基因，进行生物信息学分析。用RT-PCR检测GzCIPK7-5B基因在贵紫麦1号不同组织（根、茎、叶、籽粒）中的表达情况。运用qRT-PCR检测GzCIPK7-5B在籽粒3个时期（花后10、25、35d）的表达水平。结果表明，GzCIPK7-5B基因的开放阅读框为1296bp，编码431个氨基酸，蛋白含有丝氨酸―苏氨酸蛋白激酶家族保守结构域，具有CIPKs家族基因的特征。其编码的蛋白含有29个磷酸化位点，无跨膜结构，是一种无信号肽的不稳定亲水性核蛋白。GzCIPK7-5B基因与野生二粒小麦的TdCIPK7-5B序列相似度最高，蛋白序列同源性为100%，在根、茎、叶和籽粒中均有表达，在贵紫麦1号籽粒花青素合成3个重要时期（花后10、25、35d），花后25和35d的表达水平显著高于花后10d。

关键词: 小麦, GzCIPK7-5B基因, RT-PCR克隆, 生物信息学, 基因表达

Abstract:

CIPK is a kind of serine/threonine protein kinase specifically targeting calcineurin B similar protein in plant calcium receptors,which plays an important role in calcium ion signal transduction.To explore the function of GzCIPK7-5B gene in wheat,the GzCIPK7-5B gene from wheat variety‘Guizimai 1’was cloned by RT-PCR and analyzed by bioinformatics.RT-PCR was used to detect the expression of GzCIPK7-5B gene in different tissues (roots, stems, leaves and grains) of ‘Guizimai 1’.qRT-PCR was used to detect the expression levels of GzCIPK7-5B at three stages (10, 25, 35days post anthesis) of grain.The results showed that the length of GzCIPK7-5B open reading frame was 1296bp,encoding 431 amino acids. The protein contained a conserved domain of serine-threonine protein kinase family and had the characteristics of CIPKs family genes. The encoded protein contained 29 phosphorylation sites and had no transmembrane structure. It was an unstable hydrophilic nuclear protein without signal peptide. The sequence similarity of GzCIPK7-5B gene and TdCIPK7-5B of wild two-grain wheat was the highest, and the protein sequence homology was 100%.GzCIPK7-5B gene was expressed in roots, stems,leaves and grains. In the three important periods of anthocyanin synthesis of ‘Guizimai 1’,the expression levels of 25 and 35days post anthesis were significantly higher than that of 10days after flowering.

Key words: Wheat, GzCIPK7-5B gene, RT-PCR cloning, Bioinformatics, Gene expression

赵鹏鹏, 李鲁华, 任明见, 安畅, 洪鼎立, 李欣, 徐如宏. 小麦GzCIPK7-5B基因的生物信息学及表达分析[J]. 作物杂志, 2023 (4): 77–84

Zhao Pengpeng, Li Luhua, Ren Mingjian, An Chang, Hong Dingli, Li Xin, Xu Ruhong. Bioinformatics and Expression Analysis of GzCIPK7-5B Gene in Wheat[J]. Crops, 2023(4): 77–84

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引物名称Name of primer	引物序列Sequence of primer	用途Usage
CIPK7-5B-F	CCTCTAGAATGGCCGTCGCCAAGAGCA	基因克隆
CIPK7-5B-R	CCCCCGGGTCACAATTCCTCGCATCCATGCCAC	基因克隆
CIPK7-5B-F	CGTCTTCCTCCAGCTCGTCTCC	qRT-PCR和RT-PCR
CIPK7-5B-R	GATGAGGACGTTCTGCGGCTTG	qRT-PCR和RT-PCR
Actin-F	CCAAGGCGGAGTACGATGAGTCT	qRT-PCR和RT-PCR
Actin-R	TTCATACAGCAGGCAAGCACCAT	qRT-PCR和RT-PCR

在线软件Online software	用途Usage
http://web.expasy.org/prot param/	蛋白的理化性质
https://www.cbs.dtu.dk/services/SignalP-3.0/	信号肽预测
https://web.expasy.org/protscale/	亲疏水性预测
https://www.ncbi.nlm.nih.gov/Structure/cdd/wrpsb.cgi	保守结构域预测
http://www.cbs.dtu.dk/services/	磷酸化位点预测
https://www.csbio.sjtu.edu.cn/bioinf/Cell-PLoc-2/	亚细胞定位预测
https://www.csbio.sjtu.edu.cn/bioinf/Cell-PLoc-2/	二级结构预测
https://swissmodel.expasy.org/	三级结构预测
MAGE 7.0	进化树构建
DNAMAN 8.0	多重序列比对