Identification and Expression Patterns Analysis of ADH Gene Family in Salvia miltiorrhiza

doi:10.13560/j.cnki.biotech.bull.1985.2024-1233

Abstract

Abstract:

Objective Alcohol dehydrogenase (ADH) is one of the key enzymes in plant fermentation metabolism under anaerobic conditions, and plays important roles in plant resistance to waterlogging stress. This work is aimed to identify the SmADH genes from Salvia miltiorrhiza and provide a theoretical basis for the functional study of the SmADH gene family and the breeding of flood-resistant varieties. Method Bioinformatics methods were used to identify members of the gene family based on the S. miltiorrhiza genome data, and to analyze their physicochemical properties, sequence characteristics, evolutionary, and expression regulation. The RT-qPCR was then used to study the expression patterns of SmADHs under different stresses. Result A total of seven SmADH genes were identified, belonging to two subfamilies. SmADH proteins were mainly acidic proteins. The number of encoded amino acids ranged from 379 to 438, with a molecular weight of 40.79 to 46.89 kD. Except for SmADH5/6 localized in chloroplasts, other members were localized in the cytoplasm. Genetic feature analysis showed that members of the same subfamily of SmADHs possessed conserved motifs, similar intron numbers, and intron distributions. Chromosome localization revealed that SmADH1 was located on chromosome 1, SmADH2/3/4 on chromosome 2, SmADH5/6 on chromosome 6, and SmADH7 on contig030. Regulation by miRNA analysis showed that SmADH3/4/5/6 were regulated by miRNA family members such as smi-miR167, smi-miRN5 and smi-miR482. Promoter cis-acting element analysis showed that the promoter regions of SmADHs contained a large number of stress-responsive and hormone-responsive elements. Transcriptome data and qRT-PCR analysis indicated that SmADH2/5/6 had the highest expressions in S. miltiorrhiza roots. Under waterlogging stress, SmADH2/6 expression was rapidly upregulated. After salt stress, the expression of SmADH2/6 was significantly upregulated, while the expression of SmADH5 was inhibited. The expression of SmADH2/5/6 was significantly upregulated after PEG treatment, while the expression of SmADH6 was induced by GA3. Conclusion SmADH2/5/6, which are highly expressed in the roots of S. miltiorrhiza, play a role under various stresses, and SmADH2/6 play a particularly important role in resisting waterlogging stress.

Key words: Salvia miltiorrhiza, ADH, expression analysis, waterlogging stress, gene family

HUA Wen-ping, LIU Fei, HAO Jia-xin, CHEN Chen. Identification and Expression Patterns Analysis of ADH Gene Family in Salvia miltiorrhiza[J]. Biotechnology Bulletin, 2025, 41(8): 211-219.

Figures/Tables 8

References 31

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基因Gene	引物序列 Primer sequence （5′-3′）
SmActin	F-AGGAACCACCGATCCAGACA
（DQ243702）	R-GGTGCCCTGAGGTCCTGTT
SmADH2	F-GAACCCAAAAGACTACAGCAAGCC R-ACGCCAACAAGAACAGCCACTC
SmADH5	F-GTGTGATGCCAGCAAACGGATT R-TGACGGCGACGGAGTTACCA
SmADH6	F-ATGGGAGGTCTTGCTGAGTTTTGT R-CAATGGCATCACCAGGACGC

基因Gene	引物序列 Primer sequence （5′-3′）
SmActin	F-AGGAACCACCGATCCAGACA
（DQ243702）	R-GGTGCCCTGAGGTCCTGTT
SmADH2	F-GAACCCAAAAGACTACAGCAAGCC R-ACGCCAACAAGAACAGCCACTC
SmADH5	F-GTGTGATGCCAGCAAACGGATT R-TGACGGCGACGGAGTTACCA
SmADH6	F-ATGGGAGGTCTTGCTGAGTTTTGT R-CAATGGCATCACCAGGACGC

基因Gene	基长度 Gene length （bp）	编码序列长度CDs length （bp）	内含子数目 Number of introns	氨基酸数目Number of amino acids	理论等电点pI	分子量Molecular weight/kD	平均亲水性指数 GRAVY	亚细胞定位Subcellular localization
SmADH1	2 392	1 149	9	382	6.38	41.32	-0.014	Cytoplasm
SmADH2	2 279	1 140	8	379	5.98	40.87	0.101	Cytoplasm
SmADH3	8 279	1 149	8	382	6.87	41.55	0.259	Cytoplasm
SmADH4	2 824	1 167	9	388	5.75	41.83	0.137	Cytoplasm
SmADH5	4 670	1 299	7	432	8.67	46.20	-0.059	Chloroplasts
SmADH6	4 849	1 317	7	438	8.74	46.89	-0.056	Chloroplasts
SmADH7	3 827	1 140	8	379	6.37	40.79	0.063	Cytoplasm

基因Gene	基长度 Gene length （bp）	编码序列长度CDs length （bp）	内含子数目 Number of introns	氨基酸数目Number of amino acids	理论等电点pI	分子量Molecular weight/kD	平均亲水性指数 GRAVY	亚细胞定位Subcellular localization
SmADH1	2 392	1 149	9	382	6.38	41.32	-0.014	Cytoplasm
SmADH2	2 279	1 140	8	379	5.98	40.87	0.101	Cytoplasm
SmADH3	8 279	1 149	8	382	6.87	41.55	0.259	Cytoplasm
SmADH4	2 824	1 167	9	388	5.75	41.83	0.137	Cytoplasm
SmADH5	4 670	1 299	7	432	8.67	46.20	-0.059	Chloroplasts
SmADH6	4 849	1 317	7	438	8.74	46.89	-0.056	Chloroplasts
SmADH7	3 827	1 140	8	379	6.37	40.79	0.063	Cytoplasm

miRNA	靶基因 Target gene	期望值Expectation	靶位点Target site （bp）	抑制方式 Inhibition way
smi-miR167a	SmADH3	4	60-80	剪切
smi-miR167b	SmADH3	4	60-80	剪切
smi-miR167c	SmADH3	4	60-80	剪切
smi-miR167e	SmADH3	4	60-80	剪切
smi-miRN43	SmADH4	4.5	1 079-1 099	剪切
smi-miR482c	SmADH5	5	601-622	翻译
smi-miR482d	SmADH5	5	601-622	翻译
smi-miRN18	SmADH6	3.5	377-398	翻译
smi-miR482c	SmADH6	5	619-640	翻译
smi-miR482d	SmADH6	5	619-640	翻译
smi-miRN5a	SmADH7	4.5	5-25	翻译
smi-miRN5b	SmADH7	4.5	5-25	剪切
smi-miR530a	SmADH7	5	535-555	剪切