大豆LAZ1基因家族鉴定与GmLAZ1-9基因的功能研究

doi:10.3969/j.issn.1004-1524.2021.04.03

浙江农业学报 ›› 2021, Vol. 33 ›› Issue (4): 586-594.DOI: 10.3969/j.issn.1004-1524.2021.04.03

大豆LAZ1基因家族鉴定与GmLAZ1-9基因的功能研究

杨昕霞^a(), 张斌^b^,^*()

a. 后勤处湖南科技学院
b.化学与生物工程学院,湖南永州 425199

收稿日期:2020-10-11 出版日期:2021-04-25 发布日期:2021-04-25
通讯作者: 张斌
作者简介:*张斌,E-mail:zhangbin27104@163.com
杨昕霞(1982—),女,湖南永州人,学士,副教授,研究方向为植物资源利用。E-mail:76406391@qq.com
基金资助:
湖南省自然科学基金(2020JJ4030)

Identification of soybean LAZ1 gene family and functional analysis of GmLAZ1-9

YANG Xinxia^a(), ZHANG Bin^b^,^*()

a. Department of Logistics
b. College of Chemistry and Bioengineering, Hunan University of Science and Engineering, Yongzhou 425199, China

Received:2020-10-11 Online:2021-04-25 Published:2021-04-25
Contact: ZHANG Bin

摘要/Abstract

摘要：

通过在大豆基因组数据库中比对,获得大豆LAZ1基因家族成员,开展生物信息学分析与表达模式检测,并利用转基因拟南芥进行基因功能验证。结果显示:共鉴定到17个大豆LAZ1基因,即GmLAZ1-1~ GmLAZ1-17。这17个GmLAZ1基因不均匀地分布在大豆的12条染色体上,在进化树上可分为3个亚家族,同一亚家族成员具有类似的基因和蛋白结构。顺式作用元件预测结果表明,大豆LAZ1家族基因启动子上存在很多与非生物胁迫相关的元件,如ABRE、ARE、LTR、G-box等。半定量PCR检测结果显示,GmLAZ1-5、GmLAZ1-9、GmLAZ1-11和GmLAZ1-13在盐处理后的大豆幼苗中表达上调。在拟南芥中过表达GmLAZ1-9基因增强了转基因植株的耐盐性。这些发现可为进一步探究其他LAZ1基因的功能与分子机制提供依据。

关键词: 大豆, LAZ1家族, 盐胁迫, 基因表达

Abstract:

In the present study, the BLAST search was performed in the soybean genome database and LAZ1 gene family members were obtained. Bioinformatics analysis and expression pattern detection were performed, and transgenic Arabidopsis was used to verify gene function. It was shown that a total of 17 soybean LAZ1 genes were identified, namely GmLAZ1-1 to GmLAZ1-17. All of the GmLAZ1 genes were unevenly distributed on 12 chromosomes. The LAZ1 family members in soybean were phylogenetically divided into three evolutionary clades, and the members in the same clades shared similar gene and protein structures. The analysis of cis-regulatory elements indicated that multiple abiotic stress-related cis-acting elements presented in the promoters of soybean LAZ1 genes, such as ABRE, ARE, LTR, G-box, etc. Semi-quantitative PCR results showed that the expression of GmLAZ1-5, GmLAZ1-9, GmLAZ1-11, and GmLAZ1-13 was up-regulated after salt treatment. Moreover, GmLAZ1-9 enhanced salinity tolerance in transgenic Arabidopsis, which provided basis for further exploration of the functions and molecular mechanisms of other LAZ1 genes.

Key words: soybean, LAZ1 family, salt stress, gene expression

中图分类号:

S565.1
Q78

杨昕霞, 张斌. 大豆LAZ1基因家族鉴定与GmLAZ1-9基因的功能研究[J]. 浙江农业学报, 2021, 33(4): 586-594.

YANG Xinxia, ZHANG Bin. Identification of soybean LAZ1 gene family and functional analysis of GmLAZ1-9[J]. Acta Agriculturae Zhejiangensis, 2021, 33(4): 586-594.

图/表 5

表1 大豆LAZ1家族基因成员的理化性质

Table 1 Physical and chemical parameters of genes in soybean LAZ1 family

基因名 Name	基因编号 Gene ID	基因长度 Gene length/bp	氨基酸长度 Peptide residues/aa	相对分子质量 Molecular weight/ku	pI	不稳定系数 Instability index
GmLAZ1-1	Glyma.03G226700	5 087	421	47.17	6.78	44.35
GmLAZ1-2	Glyma.04G001400	5 252	486	54.56	8.75	44.04
GmLAZ1-3	Glyma.04G240300	5 229	288	32.65	5.85	49.15
GmLAZ1-4	Glyma.06G000800	5 037	492	55.03	8.86	41.80
GmLAZ1-5	Glyma.06G123100	5 818	485	55.25	6.77	47.60
GmLAZ1-6	Glyma.06G253700	6 131	304	35.01	8.79	26.55
GmLAZ1-7	Glyma.09G071000	7 053	440	49.44	4.35	42.21
GmLAZ1-8	Glyma.10G141000	7 265	418	46.92	8.23	41.65
GmLAZ1-9	Glyma.12G147700	9 833	287	33.18	8.87	31.34
GmLAZ1-10	Glyma.13G038900	7 402	484	54.98	5.56	46.63
GmLAZ1-11	Glyma.13G245500	2 971	296	34.07	8.80	43.68
GmLAZ1-12	Glyma.14G122700	8 650	484	55.12	5.99	44.47
GmLAZ1-13	Glyma.15G068200	2 547	296	34.13	8.59	45.68
GmLAZ1-14	Glyma.15G178900	4 890	397	44.60	8.84	32.32
GmLAZ1-15	Glyma.17G156700	4 054	384	44.25	9.56	37.51
GmLAZ1-16	Glyma.19G223700	4 841	419	46.68	6.31	41.23
GmLAZ1-17	Glyma.20G089700	3 907	418	46.77	8.41	40.32

图1 大豆LAZ1家族成员的生物信息学分析 A,大豆LAZ1家族基因的染色体定位;B,大豆和其他物种(拟南芥、水稻和玉米)LAZ1家族成员的系统进化树;C,大豆LAZ1家族蛋白和基因结构,及氨基酸保守基序分析。

Fig.1 Bioinformatics analysis of soybean LAZ1 family members A, Chromosomal location of soybean LAZ1 family genes; B, Phylogenetic tree of members of the LAZ1 family of soybeans and other species (Arabidopsis, rice, and maize); C, Analysis of gene structure and amino acid conservative motifs of soybean LAZ1 family members.

图2 大豆LAZ1家族基因在不同组织中的表达模式右侧色彩刻度标尺代表基因表达量的高低,从灰到红色表示表达量数值从低到高。

Fig.2 Expression patterns of soybean LAZ1 genes in different tissues The color scale on the right represents the level of gene expression. From gray to red, the value of expression increases from low to high.

图3 大豆LAZ1家族基因启动子区的顺式作用元件与LAZ1家族基因在盐处理条件下的表达模式 A,大豆LAZ1家族基因启动子区与非生物胁迫响应相关的顺式作用元件;B,利用半定量PCR检测LAZ1家族基因在150 mmol·L-1 NaCl处理0、3、6、12 h时的表达情况。

Fig.3 Cis-acting element in soybean LAZ1 gene promoters and expression patterns of LAZ1 genes under salt treatment A, Abiotic stress-related cis-acting elements in the promoters of soybean LAZ1 genes; B, Expression of LAZ1 genes at 0, 3, 6, 12 h after treatment with 150 mmol·L-1 NaCl based on semi-quantitative PCR test.

图4 过表达大豆GmLAZ1-9基因增强转基因拟南芥的耐盐性野生型拟南芥WT和转基因材料OEGmLAZ1-9在1/2MS培养基和1/2MS+200 mmol·L-1 NaCl培养基上萌发7 d的表型(A)和萌发率统计(B);生长4周的拟南芥(WT和OEGmLAZ1-9)在正常生长和150 mmol·L-1 NaCl溶液处理7 d的表型(C)、叶片相对含水量(D)、MDA含量(E)、脯氨酸含量(F)。D~F图中柱上标“*”的表示不同材料间差异显著(P<0.05)。

Fig.4 Overexpression of soybean GmLAZ1-9 gene enhanced salt tolerance of transgenic Arabidopsis Phenotype (A) and germination rate (B) of the wild-type Arabidopsis WT and the transgenic material OEGmLAZ1-9 seeds for 7 days on 1/2MS medium and 1/2MS+200 mmol·L-1 NaCl medium; Phenotype (C), relative water content (D), and content of MDA (E) and proline (F) of 4-week-old Arabidopsis(WT and OEGmLAZ1-9) grown in normal condition and 150 mmol·L-1 NaCl solution for 7 days. Bars in D-F marked with “*” indicated significant difference between test materials at P<0.05 under the same condition.

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大豆LAZ1基因家族鉴定与GmLAZ1-9基因的功能研究

Identification of soybean LAZ1 gene family and functional analysis of GmLAZ1-9

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