大蒜叶片蜡质成分分析及蜡质缺失基因Ggl-1筛选

doi:10.13560/j.cnki.biotech.bull.1985.2025-0220

摘要/Abstract

摘要：

目的分析大蒜蜡质缺失突变体8684-gl叶表蜡质缺失成分，筛选蜡质缺失基因Ggl-1，为探讨大蒜叶片表面蜡粉的分子调控机制奠定基础，也可为大蒜抗病虫害育种及其在生产实践中的应用提供理论基础。方法以大蒜蜡质缺失突变体8684-gl和野生型8684植株叶片为研究对象，利用气相色谱-质谱联用（gas chromatograph-mass spectrometer-computer, GC-MS）分析大蒜蜡质缺失突变体8684-gl及其野生型8684叶片表面的蜡质成分，通过转录组和实时荧光定量PCR（RT-qPCR）筛选蜡质缺失基因Ggl-1。结果通过GC-MS分析发现，大蒜叶片表面蜡质包含39个成分，8684-gl突变体叶片表面蜡质缺失的主要物质为16-庚烯酮（16-hentriacontanone, C₃₁H₆₂O）。转录组分析结果显示，蜡质缺失突变体8684-gl及其野生型8684间差异表达基因主要集中在脂质转运与代谢、脂肪酸生物合成降解过程以及次生代谢物的生物合成、运输和分解代谢等过程，蜡质缺失基因Ggl-1的候选基因为Asa8G04000和Asa4G02100，RT-qPCR结果显示，候选基因表达量在蜡质缺失突变体8684-gl及其野生型8684叶片中差异显著。结论蜡质缺失突变体8684-gl叶片表面蜡质缺失主要物质为16-庚烯酮，筛选到Asa8G04000和Asa4G02100为大蒜蜡质缺失基因Ggl-1的候选基因。

关键词: 大蒜, 蜡质缺失, 气相色谱-质谱联用, 蜡质成分, 转录组测序, 实时荧光定量PCR, 候选基因

Abstract:

Objective This study is aimed to analyze the wax-deficient components in the leaves of the garlic mutant 8684-gl and identify the wax-deficient gene Ggl-1. The findings may lay foundation for exploring the molecular mechanisms underlying wax formation on garlic leaf surfaces and provide a theoretical basis for garlic pest resistance breeding and its application in agricultural production. Method The garlic wax-deficient mutant 8684-gl and its wild-type counterpart 8684 were used as research materials. Gas Chromatography-Mass Spectrometry (GC-MS) was employed to analyze the wax composition on leaf surfaces. Transcriptome sequencing was conducted to identify the Ggl-1 gene, and its expression was validated using real-time quantitative PCR (RT-qPCR). Result By GC-MS analysis the 39 wax components on the surface of garlic leaves were found, with 16-hentriacontanone (C₃₁H₆₂O) as the primary missing component in 8684-gl. Transcriptome analysis revealed that differentially expressed genes (DEGs) between 8684-gl and wild-type 8684 were mainly involved in lipid transport and metabolism, fatty acid biosynthesis and degradation, and the transport and catabolism of secondary metabolites. Two candidate genes for Ggl-1, Asa8G04000 and Asa4G02100 were identified. RT-qPCR analysis confirmed that these candidate genes presented significant different expressions between 8684-gl and its wild-type counterpart. Conclusion The primary wax-deficient component in 8684-gl was 16-hentriacontanone. The gene Asa8G04000 and Asa4G02100 are identified as candidate genes for the wax-deficient gene Ggl-1, providing insights into the genetic basis of wax biosynthesis in garlic.

Key words: garlic, wax-deficient material, gas chromatograph-mass spectrometer-computer, wax components, transcriptome sequencing, real-time fluorescence quantitative PCR, candidate genes

刘泽洲, 段乃彬, 岳丽昕, 王清华, 姚行浩, 高莉敏, 孔素萍. 大蒜叶片蜡质成分分析及蜡质缺失基因Ggl-1筛选[J]. 生物技术通报, 2025, 41(9): 219-231.

LIU Ze-zhou, DUAN Nai-bin, YUE Li-xin, WANG Qing-hua, YAO Xing-hao, GAO Li-min, KONG Su-ping. Analysis of Wax Components and Screening of Wax-deficient Gene Ggl-1 in Garlic （Allium sativum L.）[J]. Biotechnology Bulletin, 2025, 41(9): 219-231.

图/表 11

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样品名Sample	原始读段数目 Raw reads （Mb）	过滤读段数目 Clean reads （Mb）	过滤后数据量 Clean data （Gb）	Q30质量值比例Q30 rate （%）	比对率Alignment rate （%）	测序年份Sequencing batch
GL1A	45.44	42.38	6.36	92.49	93.27	2023
GL2A	45.44	42.54	6.38	92.94	93.62
GL3A	45.44	42.03	6.30	93.50	92.50
GY1A	45.44	42.74	6.41	93.17	94.06
GY2A	45.44	42.41	6.36	93.81	93.35
GY3A	45.44	42.49	6.37	93.19	93.52
Ggl-1	62.61	61.63	9.25	94.53	88.86	2022
Ggl-2	74.48	71.80	10.77	92.58	88.86
Ggl-3	73.45	70.40	10.56	92.52	88.39
Gwx-1	68.59	65.25	9.79	92.91	89.24
Gwx-2	56.61	55.72	8.36	93.85	88.93
Gwx-3	71.67	68.79	10.32	92.90	87.89

样品名Sample	原始读段数目 Raw reads （Mb）	过滤读段数目 Clean reads （Mb）	过滤后数据量 Clean data （Gb）	Q30质量值比例Q30 rate （%）	比对率Alignment rate （%）	测序年份Sequencing batch
GL1A	45.44	42.38	6.36	92.49	93.27	2023
GL2A	45.44	42.54	6.38	92.94	93.62
GL3A	45.44	42.03	6.30	93.50	92.50
GY1A	45.44	42.74	6.41	93.17	94.06
GY2A	45.44	42.41	6.36	93.81	93.35
GY3A	45.44	42.49	6.37	93.19	93.52
Ggl-1	62.61	61.63	9.25	94.53	88.86	2022
Ggl-2	74.48	71.80	10.77	92.58	88.86
Ggl-3	73.45	70.40	10.56	92.52	88.39
Gwx-1	68.59	65.25	9.79	92.91	89.24
Gwx-2	56.61	55.72	8.36	93.85	88.93
Gwx-3	71.67	68.79	10.32	92.90	87.89

基因 Gene	正向引物序列 Forward sequence （5′-3′）	反向引物序列 Reverse sequence （5′-3′）
AsACTIN	TCCTAACCGAGCGAGGCTACAT	GGAAAAGCACTTCTGGGCACC
Asa8G04000	CACTGCTGCTGTGCTTTCAG	ACTGGGACGATCTTCCTGGA
Asa4G02100	TGAGGTGATGCCAGTGAACC	TCACCCAGTCGAAATGTGCA
Asa7G01089	CGCTGGTAGTCCACACCTTT	TAACGTGTCCAAGCTCCCAC
Asa5G03876	TTGGCAAGAGTAGCAGCTCC	GATCGAATAGCGACTGGCCA
Asa7G02324	GTCCTCTGGAATTGCTGCCT	CGCCTTCATACGCCCATTTG
Asa4G04412	AGCGCTGGAGTTGGAGTTAC	ATGTTCATCACAGCCCCTGG
Asa1G04660	TGTAGGATCTCGTTCGGGGT	GACGACAGGGTTTACAGCCA
Asa1G03601	CCTGAGTTCGTCGATACCGG	TCCATAAACATGCACCCGCT
Asa1G03571	TGAAGCGTCGCAAAACTGTG	TTTCTCCGGGGCACTTCATC
Asa4G06138	GGAAGCTTCCCATGTTCGGA	ACGGACGTTGTCAAATCCGA
Asa2G01695	TTAACCGCTTGTTTGCACCG	TAGTCAACAGCCAGCATGCA
Asa0G03810	AGGGCATCCTTGGGGAAATG	TGTTCGGGACTCCATTTGCA

基因 Gene	正向引物序列 Forward sequence （5′-3′）	反向引物序列 Reverse sequence （5′-3′）
AsACTIN	TCCTAACCGAGCGAGGCTACAT	GGAAAAGCACTTCTGGGCACC
Asa8G04000	CACTGCTGCTGTGCTTTCAG	ACTGGGACGATCTTCCTGGA
Asa4G02100	TGAGGTGATGCCAGTGAACC	TCACCCAGTCGAAATGTGCA
Asa7G01089	CGCTGGTAGTCCACACCTTT	TAACGTGTCCAAGCTCCCAC
Asa5G03876	TTGGCAAGAGTAGCAGCTCC	GATCGAATAGCGACTGGCCA
Asa7G02324	GTCCTCTGGAATTGCTGCCT	CGCCTTCATACGCCCATTTG
Asa4G04412	AGCGCTGGAGTTGGAGTTAC	ATGTTCATCACAGCCCCTGG
Asa1G04660	TGTAGGATCTCGTTCGGGGT	GACGACAGGGTTTACAGCCA
Asa1G03601	CCTGAGTTCGTCGATACCGG	TCCATAAACATGCACCCGCT
Asa1G03571	TGAAGCGTCGCAAAACTGTG	TTTCTCCGGGGCACTTCATC
Asa4G06138	GGAAGCTTCCCATGTTCGGA	ACGGACGTTGTCAAATCCGA
Asa2G01695	TTAACCGCTTGTTTGCACCG	TAGTCAACAGCCAGCATGCA
Asa0G03810	AGGGCATCCTTGGGGAAATG	TGTTCGGGACTCCATTTGCA

材料Material	株高Plant height （cm）	假茎高Pseudostem height （cm）	叶长Leaf length （cm）	叶宽Leaf width （cm）	茎粗Stem thick （cm）
8684	92.40±3.78	36.95±1.86	58.70±2.70	3.64±0.23	18.08±1.59
8684-gl	94.55±2.83	37.75±1.86	59.60±2.50	3.46±0.20	17.21±1.01