高抗旱性二倍体马铃薯材料转录组分析及抗旱基因的初步挖掘

doi:10.7668/hbnxb.20193446

摘要/Abstract

摘要：

为探究不同干旱胁迫时间处理下,二倍体马铃薯材料对干旱胁迫响应的分子机制,挖掘抗旱相关基因。以高抗旱二倍体资源A90为试验材料,利用转录组测序技术对PEG-6000胁迫下不同时间的差异表达基因进行分析,通过GO富集、KEGG通路分析与转录因子预测参与马铃薯干旱胁迫响应的差异表达基因,初步挖掘抗旱调控关键基因;并通过RT-qPCR对3个抗旱候选基因进行逆境胁迫表达分析。结果表明:A90在20%PEG-6000胁迫处理3,6,24 h与对照相比,共有2 519个差异表达基因,这些基因主要富集在植物激素信号转导、谷胱甘肽代谢、苯丙烷生物合成、戊糖和葡萄糖醛酸相互转化等抗旱相关过程中。并且显著富集在通路上的340个共差异表达基因中有53个基因注释到了RLKs、AP2/ERF和Tify等15个转录因子家族中,其中StST/K1、StERF1和StTify1的表达量较高,有可能是抗旱调控关键基因。基因StST/K1和StERF1受到干旱和低温胁迫主要在材料的根和叶中上调表达,基因StERF1在盐胁迫下在材料的根、茎和叶中均上调表达;基因StTify1在受到干旱在材料的茎和叶中上调表达,受到盐胁迫在材料的根、茎和叶中均上调表达,但在低温胁迫下只在材料的茎中下调表达。由此说明,筛选出的3个基因均能够对干旱、盐和低温胁迫产生响应,可为今后马铃薯抗旱分子育种中相关抗旱候选基因研究提供一定的理论基础。

关键词: 马铃薯, 二倍体, 干旱胁迫, 转录组测序, 抗旱候选基因

Abstract:

In order to explore the molecular mechanism of diploid potato material response to drought stress under different drought stress time treatment,and explore drought resistance related genes.The high drought-resistant diploid resource A90 was used as experimental material,and transcriptome sequencing technology was used to analyze the differentially expressed genes under PEG-6000 stress at different times.Differentially expressed genes involved in drought stress response of potato were predicted by GO enrichment,KEGG pathway analysis and transcription factors,and key genes of drought resistance regulation were preliminarily explored.The expression of three candidate genes was analyzed by RT-qPCR.The results showed that there were 2 519 differentially expressed genes in the A90 treated with 20% PEG-6000 for 3,6,24 h compared with the control,and these genes were mainly enriched in plant hormone signal transduction,glutathione metabolism,phenylpropanoid biosynthesis,pentose and glucuronate interconversions and other drought-related processes.In addition,among 340 codifferentially expressed genes significantly enriched in the pathway,53 genes were annotated to 15 transcription factor families including RLKs,AP2/ERF and Tify.Among which StST/K1,StERF1 and StTify1 had higher expression levels,which might be key genes in drought resistance regulation.StST/K1 and StERF1 were mainly up-regulated in roots and leaves under drought and low temperature stress,StERF1 was up-regulated in roots,stems and leaves under salt stress.StTify1 was up-regulated in stems and leaves of the material under drought,and it was up-regulated in roots,stems and leaves under salt stress.But it was down-regulated only in stems of the material under low temperature stress.These results indicate that the three selected genes can respond to drought,salt and low temperature stress,it can provide a theoretical basis for study on drought resistance candidate genes in potato molecular breeding in the future.

Key words: Potato, Diploid, Drought stress, Transcriptome sequencing, Candidate genes for drought resistance

季香林, 张丽莉, 甘珊, 石瑛. 高抗旱性二倍体马铃薯材料转录组分析及抗旱基因的初步挖掘[J]. 华北农学报, 2023, 38(1): 63-73. doi: 10.7668/hbnxb.20193446.

JI Xianglin, ZHANG Lili, GAN Shan, SHI Ying. Transcriptome Analysis of Diploid Potato Material with High Drought Tolerance and Preliminary Excavation of Drought Resistance Genes[J]. Acta Agriculturae Boreali-Sinica, 2023, 38(1): 63-73. doi: 10.7668/hbnxb.20193446.

http://www.hbnxb.net/CN/Y2023/V38/I1/63

图/表 13

参考文献 32

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基因名称 Gene name	正向引物(5'—3') Forward primer(5'—3')	反向引物(5'—3') Reverse primer(5'—3')
Soltu.DM.07G025300	TTCCCTTTGTCACTTCAC	CTCCTTCTTGTCCCTTTC
Soltu.DM.01G043800	TTACATCCCCAAAACCCA	TTTCCCTCCACACCTCAT
Soltu.DM.08G029550	TCCATCATTCAATCTCAA	ACAAAGTTACTCCACCCT
Soltu.DM.12G007690	TGGAATGCGAAAGATAGG	GATGGGAGGAGGAGTGAG
Elfα	GATGTTGTGCCAAAGGATGT	AACTTGGTCAATGCGAGA

基因名称 Gene name	正向引物(5'—3') Forward primer(5'—3')	反向引物(5'—3') Reverse primer(5'—3')
Soltu.DM.07G025300	TTCCCTTTGTCACTTCAC	CTCCTTCTTGTCCCTTTC
Soltu.DM.01G043800	TTACATCCCCAAAACCCA	TTTCCCTCCACACCTCAT
Soltu.DM.08G029550	TCCATCATTCAATCTCAA	ACAAAGTTACTCCACCCT
Soltu.DM.12G007690	TGGAATGCGAAAGATAGG	GATGGGAGGAGGAGTGAG
Elfα	GATGTTGTGCCAAAGGATGT	AACTTGGTCAATGCGAGA

基因名称 Gene name	正向引物(5'—3') Forward primer(5'—3')	反向引物(5'—3') Reverse primer(5'—3')
Soltu.DM.03G029730	GAGATACCCTGGAGAGTG	TGAGTGAAAAATGACGAA
Soltu.DM.05G020900	CAACAACATCAAAAGCAA	GAGAAGAGACCAAGGACC
Soltu.DM.09G021200	ATTTGGGTGCTGAGTATT	CACGTTGTTGGTGAGAGA
Soltu.DM.08G007150	AACTCACTCTTTCGCTTT	TTCTCCTCCATTCCTCTA
Elfα	GATGTTGTGCCAAAGGATGT	AACTTGGTCAATGCGAGA

基因名称 Gene name	正向引物(5'—3') Forward primer(5'—3')	反向引物(5'—3') Reverse primer(5'—3')
Soltu.DM.03G029730	GAGATACCCTGGAGAGTG	TGAGTGAAAAATGACGAA
Soltu.DM.05G020900	CAACAACATCAAAAGCAA	GAGAAGAGACCAAGGACC
Soltu.DM.09G021200	ATTTGGGTGCTGAGTATT	CACGTTGTTGGTGAGAGA
Soltu.DM.08G007150	AACTCACTCTTTCGCTTT	TTCTCCTCCATTCCTCTA
Elfα	GATGTTGTGCCAAAGGATGT	AACTTGGTCAATGCGAGA

样品名称 Sample name	有效数据 Clean reads	总映射 Total mapped	总映射率/% Total mapped rate	唯一映射 Uniquely mapped	唯一映射率/% Uniquely mapped rate	GC含量/% GC content	Q30/%
T0A90a	43 435 584	40 013 630	92.12	38 912 643	89.59	42.80	93.98
T0A90b	46 543 524	42 716 607	91.78	41 533 558	89.24	42.69	93.95
T0A90c	41 533 642	38 276 615	92.16	37 266 032	89.72	42.75	94.80
T3A90a	43 204 066	39 417 930	91.24	38 283 237	88.61	42.60	93.94
T3A90b	45 816 482	41 000 567	89.49	39 438 548	86.08	43.04	93.64
T3A90c	46 049 908	41 805 803	90.78	40 513 499	87.98	42.78	93.84
T6A90a	44 475 160	40 591 004	91.27	39 383 761	88.55	42.49	94.41
T6A90b	43 682 456	39 550 185	90.54	38 338 301	87.77	42.70	93.65
T6A90c	43 481 150	39 499 021	90.84	38 294 871	88.07	42.62	93.62
T24A90a	42 662 576	38 618 685	90.52	37 448 658	87.78	42.50	94.70
T24A90b	43 157 618	37 343 483	86.53	35 182 612	81.52	43.88	94.14
T24A90c	47 315 172	43 117 670	91.13	41 838 025	88.42	42.58	94.73

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