谷子B3转录因子可变剪切体分析

doi:10.3969/j.issn.1004-1524.20231216

浙江农业学报 ›› 2024, Vol. 36 ›› Issue (9): 1969-1976.DOI: 10.3969/j.issn.1004-1524.20231216

谷子B3转录因子可变剪切体分析

刘辉¹(), 王晓蒙¹, 闫留延¹, 王永芳², 杨朋娟¹, 龚珂珂¹, 李兴杰¹, 董志平², 贾小平¹^,^*()

1.河南科技大学农学院,河南洛阳 471023
2.河北省农林科学院谷子研究所,国家谷子改良中心,河北石家庄 050035

收稿日期:2023-10-31 出版日期:2024-09-25 发布日期:2024-09-30
作者简介:*贾小平,E-mail:jiaxiaoping2007@163.com
刘辉(1998—),男,江苏盐城人,硕士研究生,研究方向为谷子遗传育种。E-mail:hliu0921@163.com
通讯作者: *贾小平,E-mail:jiaxiaoping2007@163.com
基金资助:
国家自然科学基金(31471569)

Analysis of the B3 transcription factor alternative splicer in foxtail millet (Setaria italica L.)

LIU Hui¹(), WANG Xiaomeng¹, YAN Liuyan¹, WANG Yongfang², YANG Pengjuan¹, GONG Keke¹, LI Xingjie¹, DONG Zhiping², JIA Xiaoping¹^,^*()

1. College of Agriculture, Henan University of Science and Technology, Luoyang 471023, Henan, China
2. National Millet Improvement Center, Institute of Millet, Hebei Academy of Agriculture and Forestry Sciences, Shijiazhuang 050035, China

Received:2023-10-31 Online:2024-09-25 Published:2024-09-30

摘要/Abstract

摘要：

通过分析不同谷子品种B3转录因子基因存在的可变剪切类型,鉴定出具有功能活性的可变剪切体,并分析这些可变剪切体与谷子抽穗期是否具有相关性,为深入揭示B3转录因子可变剪切对谷子生长发育的调控作用提供依据。选择12份不同抽穗期的谷子材料,利用反转录PCR(reverse transcription PCR, RT-PCR)技术克隆了一个REM家族的B3转录因子基因,通过序列比对鉴定该基因存在的可变剪切体类型,分析这些可变剪切体的活性及其与抽穗期的关系。结果表明,B3转录因子基因在这12个谷子品种间共产生了8种可变剪切体,其中可变剪切体Ⅴ包含的品种最多(4个),其次为可变剪切体Ⅳ(2个),其余6种可变剪切体均只包含1个品种。通过可变剪切,12个品种的B3转录因子基因共可编码10种蛋白质,其中5种蛋白质缺失B3 DNA结合域;Ⅱ型蛋白质具有缩短谷子抽穗期的功能,Ⅵ型蛋白质具有延长谷子抽穗期的功能,且这种作用不受地理环境影响。以上结果表明,B3转录因子通过可变剪切参与谷子抽穗期的调控。

关键词: 谷子, B3转录因子, 可变剪切, 抽穗期, 地理环境

Abstract:

The alternative splicing types of B3 transcriptional factor gene in different millet varieties were analyzed to identify the splicing variants with functional activity. Furthermore, the correlation between these splicing variants and heading stage was analyzed, aiming at providing evidence for further revealing the regulatory effects of B3 transcription factor on growth and development of millet by alternative splicing. Totally 12 millet varieties with different heading stages were selected to clone B3 transcriptional factor gene belonging to REM family by reverse transcription PCR (RT-PCR). The alternative splicing types of this gene were identified by multiple sequence alignment, and the activity of the splicing variants, their association with heading stage were analyzed. The results showed that a total of 8 splicing variants of B3 transcriptional factor gene were produced among the 12 millet varieties, of which splicing variant Ⅴ contained the most varieties (4), followed by splicing variant Ⅳ (2), and the remaining 6 splicing variants each contained only 1 variety. By alternative splicing, the B3 transcriptional factor gene of 12 varieties encoded 10 types of proteins, of which 5 types lacked the B3 DNA binding domain, and type Ⅱ protein shortened the heading stage of millet, while type Ⅵ protein prolonged the heading stage of millet, which was not affected by geographical environments. The above results showed that B3 transcription factor participated in the regulation of heading stage through alternative splicing in millet.

Key words: foxtail millet, B3 transcription factor, alternative splicing, heading stage, geographic environment

中图分类号:

S515

刘辉, 王晓蒙, 闫留延, 王永芳, 杨朋娟, 龚珂珂, 李兴杰, 董志平, 贾小平. 谷子B3转录因子可变剪切体分析[J]. 浙江农业学报, 2024, 36(9): 1969-1976.

LIU Hui, WANG Xiaomeng, YAN Liuyan, WANG Yongfang, YANG Pengjuan, GONG Keke, LI Xingjie, DONG Zhiping, JIA Xiaoping. Analysis of the B3 transcription factor alternative splicer in foxtail millet (Setaria italica L.)[J]. Acta Agriculturae Zhejiangensis, 2024, 36(9): 1969-1976.

图/表 6

表1 特异性引物信息

Table 1 Information of specific primers

引物名称 Primer name	正向引物(5’→3’) Forward primer (5’→3’)	反向引物(5’→3’) Reverse primer (5’→3’)	片段大小 Fragment size/bp
引物1 Primer 1	GTGAGGGAGGGAGAAATCG	GCTAAGTAAGTCAGGCTGAAACA	907
引物2 Primer 2	GAGAAATCGAGGGAGAAAAT	CTACCTTCCTGCTAAGACTGC	707

图1 不同品种谷子总RNA电泳结果 M,DL 2 000 DNA marker;1~12分别为郑8041、安丰、沙湾谷子、骨绿早1、大头糯、小早谷、金德、大王国、呼和浩特大毛谷、安5424、豫谷13号、糟皮一把奇。下同。

Fig.1 Electrophoresis results of total RNA from different foxtail millet varieties M, DL 2 000 DNA marker; 1-12 were Zheng 8041, Anfeng, Shawan Guzi, Gulyu Zao 1, Datou Nuo, Xiaozao Gu, Jinde, Da Kingdom, Hohhot Damao Gu, An 5424, Yugu 13, and Zaopiyibaqi, respectively. The same as below.

图2 不同谷子品种B3转录因子基因的PCR扩增产物电泳结果

Fig.2 Electrophoresis results of PCR amplification products of B3 transcription factor genes from different foxtail millet varieties

图3 十二份谷子品种的B3转录因子基因cDNA序列比对结果 Ⅰ~Ⅶ表示不同可变剪接体类型。图右边编号1~12依次对应谷子品种为郑8041、安丰、沙湾谷子、骨绿早1、大头糯、小早谷、金德、大王国、呼和浩特大毛谷、安5424、豫谷13号、糟皮一把奇。

Fig.3 cDNA sequences alignment result of B3 transcriptional factor gene in 12 millet varieties Ⅰ-Ⅶ represented different types of variable spliceosomes. The numbers 1-12 on the right correspond to the millet varieties Zheng 8041, Anfeng, Shawanguzi, Gulyuzao 1, Datounuo, Xiaozaogu, Jinde, Da Kingdom, Huhehaotedamaogu, An 5424, Yugu 13, and Zaopiyibaqi.

图4 十二份谷子品种B3转录因子蛋白质序列比对结果图左边编号1~12依次对应谷子品种为郑8041、安丰、沙湾谷子、骨绿早1、大头糯、小早谷、金德、大王国、呼和浩特大毛谷、安5424、豫谷13号、糟皮一把奇。

Fig.4 Protein sequences alignment result of B3 transcriptional factor in 12 millet varieties The numbers 1-12 on the left correspond to the millet varieties Zheng 8041, Anfeng, Shawanguzi, Gulyuzao 1, Datounuo, Xiaozaogu, Jinde, Da Kingdom, Huhehaotedamaogu, An 5424, Yugu 13, and Zaopiyibaqi.

图5 不同B3蛋白质类型谷子品种的抽穗日数

Fig.5 Heading days among millet varities with different B3 protein types

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谷子B3转录因子可变剪切体分析

Analysis of the B3 transcription factor alternative splicer in foxtail millet (Setaria italica L.)

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