中国农业科学 ›› 2022, Vol. 55 ›› Issue (14): 2797-2811.doi: 10.3864/j.issn.0578-1752.2022.14.010
收稿日期:
2021-11-01
接受日期:
2021-12-31
出版日期:
2022-07-16
发布日期:
2022-07-26
通讯作者:
王海波
作者简介:
冀晓昊,Tel:13610890936;E-mail: 基金资助:
JI XiaoHao(),LIU FengZhi,WANG BaoLiang,LIU PeiPei,WANG HaiBo(
)
Received:
2021-11-01
Accepted:
2021-12-31
Online:
2022-07-16
Published:
2022-07-26
Contact:
HaiBo WANG
摘要:
【目的】开发适用于葡萄香气表型鉴定的分子标记,为葡萄分子辅助育种提供理论依据。【方法】采用固相微萃取结合气相质谱法对45个葡萄品种进行香气组分和含量测定,并分别采用Sanger测序和扩增子测序的方法对葡萄醇酰基转移酶编码基因(VvAAT)结构变异和SNP变异进行分析。【结果】45个葡萄品种中共发现65种香气组分,可以分成酯类、醇类、萜类和醛类4种类型,其中酯类香气含量表现出显著的品种差异性,‘巨峰’等20个葡萄品种酯类香气含量丰富,而‘87-1’等25个葡萄品种检测不到酯类香气;VvAAT共发现了5种结构变异类型,类型I、II、IV和V由于过早终止密码子或片段插入变异,不能正确翻译,仅类型III可以正常翻译,根据其氨基酸序列系统进化树分析结果,又可以分成III.1和III.2两种类型,结合香气测定数据,III.1为功能型,其余均为非功能型;扩增子测序及生信分析发现了8个位于外显子区域的SNP位点(T32C、A69T、G436C、A1247G、A1818T、G1929T、A1959G和C1975G),均导致了编码氨基酸的突变,可以准确区分酯香型品种和非酯香型品种,准确率为97.8%。【结论】葡萄VvAAT位点存在丰富的遗传变异,包括基因结构变异和SNP变异;位于编码区的8个SNP位点能够准确判定葡萄酯类香气表型,可以应用于葡萄分子辅助育种。
冀晓昊,刘凤之,王宝亮,刘培培,王海波. 葡萄醇酰基转移酶编码基因遗传变异研究[J]. 中国农业科学, 2022, 55(14): 2797-2811.
JI XiaoHao,LIU FengZhi,WANG BaoLiang,LIU PeiPei,WANG HaiBo. Genetic Variation of Alcohol Acyltransferase Encoding Gene in Grape[J]. Scientia Agricultura Sinica, 2022, 55(14): 2797-2811.
表1
45个葡萄品种香气组分和含量统计"
化合物类型 Type of compound | 种类 Number | 变异范围 Variation range (ng∙g-1) | 平均值 Mean (ng∙g-1) | 方差 Variance |
---|---|---|---|---|
酯类Esters | 22 | 0—3238.77 | 465.67 | 777.60 |
醇类Alcohols | 2 | 0—164.84 | 12.58 | 31.91 |
醛类Aldehydes | 12 | 642.80—5485.97 | 1890.40 | 1171.33 |
萜类Terpenoids | 29 | 0—8548.37 | 471.41 | 1601.95 |
总计 Total | 65 | 800.43—11387.75 | 2835.44 | 2381.58 |
表2
酯香型和非酯香型葡萄品种分类"
酯香型品种 Varieties rich in volatile esters | 非酯香型品种 Varieties poor in volatile esters |
---|---|
红香蕉 Hongxiangjiao (6, 244.27) | 8611 (0, 0) |
高砂 Gaosha (9, 278.59) | 累克基特 Leikejite (0, 0) |
秦龙大穗 Qinlongdasui (8, 220.90) | 蓓蕾玫瑰 Muscat Bailey (0, 0) |
黑蜜 Heimi (9, 199.58) | 克瑞森无核 Crimson_Seedless (0, 0) |
秋蜜 Qiumi (13, 402.83) | 8612 (0, 0) |
琥珀 Hupo (9, 330.06) | 齐查卡普列 Qichakapulie (0, 0) |
红丹 Hongdan (16, 920.17) | 牧丹红 Mudanhong (0, 0) |
火星无核 Mars_Seedless (9, 408.72) | 红意大利Red_Italian (0, 0) |
着色香 Zhuosexiang (6, 543.78) | 玫瑰花 Meiguihua (0, 0) |
黑虎香 Heihuxiang (14, 591.16) | 泽玉 Zeyu (0, 0) |
藤稔 Fujiminori (13, 701.42) | 巧保2号 Qiaobao2 (0, 0) |
红富士 BenniFuji (10, 723.95) | 红巴拉多蒂 Hongbaladuodi (0, 0) |
井川1014 Jingchun1014 (9, 800.74) | 乌兹别克玫瑰 Ojin_Kokn (0, 0) |
巨峰 Kyoho (15, 1453.39) | 阿登纳玫瑰 Muscat_Adda (0, 0) |
尼加拉 Nigara (13, 1697.20) | 228 (0, 0) |
奥利文 Irsay_Oliver (7, 1708.50) | 里扎马特 Rizamat (0, 0) |
安尼斯基 Ayaneskeal (9, 1920.01) | 香槟 Champion (0, 0) |
吉香 Jixiang (9, 2200.84) | 京秀 Jingxiu (0, 0) |
紫早 Zizao (9, 2370.34) | 索罗门 Madeleine_Salomon (0, 0) |
早巨选 Zaojuxuan (19, 3238.77) | 郑州早红 Zhengzhouzaohong (0, 0) |
87-1 (0, 0) | |
晚17 Wan17 (0, 0) | |
香妃 Xiangfei (0, 0) | |
京早晶 Jingzaojing (0, 0) | |
阳光玫瑰 Shine_Musca (0, 0) |
表3
27个葡萄品种VvAAT基因型统计表"
品种 Variety | 基因名称 Gene name | NCBI注册号 GenBank accession number | 基因结构类型 Gene structure type | 表型 Phenotype |
---|---|---|---|---|
巨峰 Kyoho | VvAAT-Kyoho-1 | OK559426 | III.2 | 酯香型 Esters-rich |
VvAAT-Kyoho-2 | OK559427 | III.1 | ||
阳光玫瑰 Shine-Muscat | VvAAT-Shine_Musca-1 | OK559428 | IV | 非酯香型 Esters-poor |
VvAAT-Shine_Musca-2 | OK559429 | V | ||
着色香 Zhuosexiang | VvAAT-Zhuosexiang-1 | OK559430 | III.1 | 酯香型 Esters-rich |
VvAAT-Zhuosexiang-2 | OK559431 | V | ||
87-1 | VvAAT-87_1 | OK559432 | III.2 | 非酯香型 Esters-poor |
晚17 Wan17 | VvAAT-Wan17 | OK559433 | II | 非酯香型 Esters-poor |
8611 | VvAAT-8611-1 | OK559434 | III.2 | 非酯香型 Esters-poor |
VvAAT-8611-2 | OK559435 | III.2 | ||
香妃 Xiangfei | VvAAT-Xiangfei | OK559436 | III.2 | 非酯香型 Esters-poor |
京早晶 Jingzaojing | VvAAT-Jingzaojing | OK559437 | III.2 | 非酯香型 Esters-poor |
郑州早红Zhengzhouzaohong | VvAAT-Zhengzhouzaohong | OK559438 | V | 非酯香型 Esters-poor |
索罗门 Madeleine_Salomon | VvAAT-Madeleine_Salomon-1 | OK559439 | III.2 | 非酯香型 Esters-poor |
VvAAT-Madeleine_Salomon-2 | OK559440 | III.2 | ||
京秀 Jingxiu | VvAAT-Jingxiu-1 | OK559441 | III.2 | 非酯香型 Esters-poor |
VvAAT-Jingxiu-2 | OK559442 | II | ||
早巨选 Zaojuxuan | VvAAT-Zaojuxuan-1 | OK559443 | III.2 | 酯香型 Esters-rich |
VvAAT-Zaojuxuan-2 | OK559444 | III.1 | ||
VvAAT-Zaojuxuan-3 | OK559445 | III.2 | ||
VvAAT-Zaojuxuan-4 | OK559446 | III.2 | ||
奥利文 Irsay_Oliver | VvAAT-Irsay_Oliver | OK559447 | III.1 | 酯香型 Esters-rich |
香槟 Champion | VvAAT-Champion | OK559448 | V | 非酯香型 Esters-poor |
紫早 Zizao | VvAAT-Zizao | OK559449 | III.1 | 酯香型 Esters-rich |
里扎马特 Rizamat | VvAAT-Rizamat-1 | OK559450 | III.2 | 非酯香型 Esters-poor |
VvAAT-Rizamat-2 | OK559451 | III.2 | ||
228 | VvAAT-228 | OK559452 | III.2 | 非酯香型 Esters-poor |
安尼斯基 Ayaneskeal | VvAAT-Ayaneskeal | OK559453 | III.1 | 酯香型 Esters-rich |
阿登纳玫瑰 Muscat_Adda | VvAAT-Muscat_Adda | OK559454 | II | 非酯香型 Esters-poor |
红富士 BenniFuji | VvAAT-BenniFuji | OK559455 | III.1 | 酯香型 Esters-rich |
吉香 Jixiang | VvAAT-Jixiang | OK559456 | III.1 | 酯香型 Esters-rich |
藤稔 Fujiminori | VvAAT-Fujiminori | OK559457 | III.1 | 酯香型 Esters-rich |
尼加拉 Nigara | VvAAT-Nigara-1 | OK559458 | III.2 | 酯香型 Esters-rich |
VvAAT-Nigara-2 | OK559459 | III.1 | ||
乌兹别克玫瑰 Ojin_Kokn | VvAAT-Ojin_Kokn | OK559460 | III.2 | 非酯香型 Esters-poor |
井川1014 Jingchun1014 | VvAAT-Jingchun1014-1 | OK559461 | III.2 | 酯香型 Esters-rich |
VvAAT-Jingchun1014-2 | OK559462 | III.1 | ||
秦龙大穗 Qinlongdasui | VvAAT-Qinlongdasui-1 | OK559463 | III.1 | 酯香型 Esters-rich |
VvAAT-Qinlongdasui-2 | OK559464 | III.2 | ||
累克基特 Leikejite | VvAAT-Leikejite-1 | OK559465 | I | 非酯香型 Esters-poor |
VvAAT-Leikejite-2 | OK559466 | II |
表5
22个葡萄品种VvAAT扩增子测序结果"
品种 Variety | Clean reads (K) | Clean bases (M) | Q20 (%) | Q30 (%) | 覆盖度 Coverage (K) |
---|---|---|---|---|---|
奥利文 Irsay_Oliver | 235.22 | 30.18 | 90.66 | 87.57 | 15.09 |
早巨选 Zaojuxuan | 407.65 | 52.72 | 90.77 | 86.72 | 26.36 |
火星无核 Mars_Seedless | 359.00 | 46.58 | 90.83 | 87.73 | 23.29 |
红香蕉 Hongxiangjiao | 445.26 | 56.83 | 90.85 | 87.80 | 28.41 |
琥珀 Hupo | 553.36 | 71.05 | 91.05 | 88.03 | 35.52 |
红富士 BenniFuji | 373.19 | 47.93 | 90.47 | 87.29 | 23.96 |
秋蜜 Qiumi | 295.73 | 37.96 | 90.76 | 87.69 | 18.98 |
巨峰 Kyoho | 379.10 | 48.84 | 90.93 | 87.88 | 24.42 |
井川1014 Jingchun1014 | 518.11 | 66.35 | 90.89 | 87.84 | 33.17 |
黑虎香 Heihuxiang | 729.92 | 95.28 | 90.87 | 87.79 | 47.64 |
黑蜜 Heimi | 601.47 | 78.07 | 91.05 | 88.01 | 39.03 |
秦龙大穗 Qinlongdasui | 693.37 | 90.00 | 90.50 | 87.34 | 45.00 |
红丹 Hongdan | 338.81 | 42.87 | 90.80 | 87.78 | 21.43 |
高砂 Gaosha | 343.88 | 43.87 | 91.00 | 88.01 | 21.93 |
红巴拉多蒂 Hongbaladuodi | 521.69 | 67.30 | 91.20 | 88.17 | 33.65 |
巧保2号 Qiaobao2 | 355.88 | 45.40 | 91.04 | 88.00 | 22.70 |
泽玉 Zeyu | 1187.71 | 155.61 | 90.83 | 87.68 | 77.80 |
玫瑰花 Meiguihua | 1771.53 | 234.87 | 90.68 | 87.46 | 117.43 |
红意大利 Red Italian | 627.85 | 79.59 | 89.91 | 86.61 | 39.79 |
牧丹红 Mudanhong | 1348.14 | 173.77 | 89.67 | 86.65 | 86.88 |
8612 | 1342.69 | 173.45 | 89.86 | 86.51 | 86.72 |
克瑞森无核 Crimson Seedless | 569.85 | 70.73 | 90.29 | 87.14 | 35.36 |
表6
一代测序数据中8个SNP位点分析"
基因名称 Gene name | SNPs | 基因结构类型 Gene structure type | |||||||
---|---|---|---|---|---|---|---|---|---|
32 | 69 | 438 | 1247 | 1818 | 1929 | 1959 | 1975 | ||
VvAAT-Kyoho-1 | T | A | G | A | A | G | A | C | III.2 |
VvAAT-Kyoho-2 | C | T | C | G | T | T | G | G | III.1 |
VvAAT-Shine_Musca-1 | T | A | G | A | A | G | A | C | IV |
VvAAT-Shine_Musca-2 | T | A | G | A | A | G | A | C | V |
VvAAT-Zhuosexiang-1 | C | T | C | G | T | T | G | G | III.1 |
VvAAT-Zhuosexiang-2 | T | A | G | A | A | G | A | C | V |
VvAAT-87_1 | T | A | G | A | A | G | A | C | III.2 |
VvAAT-Wan17 | T | A | G | A | A | G | A | C | II |
VvAAT-8611-1 | T | A | G | A | A | G | A | C | III.2 |
VvAAT-8611-2 | T | A | G | A | A | G | A | C | III.2 |
VvAAT-Xiangfei | T | A | G | A | A | G | A | C | III.2 |
VvAAT-Jingzaojing | T | A | G | A | A | G | A | C | III.2 |
VvAAT-Zhengzhouzaohong | T | A | G | A | A | G | A | C | V |
VvAAT-Madeleine_Salomon-1 | T | A | G | A | A | G | A | C | III.2 |
VvAAT-Madeleine_Salomon-2 | T | A | G | A | A | G | A | C | III.2 |
VvAAT-Jingxiu-1 | T | A | G | A | A | G | A | C | III.2 |
VvAAT-Jingxiu-2 | T | A | G | A | A | G | A | C | II |
VvAAT-Zaojuxuan-1 | T | A | G | A | A | G | A | C | III.2 |
VvAAT-Zaojuxuan-2 | C | T | C | G | T | T | G | G | III.1 |
VvAAT-Zaojuxuan-3 | T | A | G | A | A | G | A | C | III.2 |
VvAAT-Zaojuxuan-4 | T | A | G | G | T | T | G | G | III.2 |
VvAAT-Irsay_Oliver | C | T | C | G | T | T | G | G | III.1 |
VvAAT-Champion | T | A | G | A | A | G | A | C | V |
VvAAT-Zizao | C | T | C | G | T | T | G | G | III.1 |
VvAAT-Rizamat-1 | T | A | G | A | A | G | A | C | III.2 |
VvAAT-Rizamat-2 | T | A | G | A | A | G | A | C | III.2 |
VvAAT-228 | T | A | G | A | A | G | A | C | III.2 |
VvAAT-Ayaneskeal | C | T | C | G | T | T | G | G | III.1 |
VvAAT-Muscat_Adda | T | A | G | A | A | G | A | C | II |
VvAAT-BenniFuji | C | T | C | G | T | T | G | G | III.1 |
VvAAT-Jixiang | C | T | C | G | T | T | G | G | III.1 |
VvAAT-Fujiminori | C | T | C | G | T | T | G | G | III.1 |
VvAAT-Nigara-1 | T | A | G | A | A | G | A | C | III.2 |
VvAAT-Nigara-2 | C | T | C | G | T | T | G | G | III.1 |
VvAAT-Ojin_Kokn | T | A | G | A | A | G | A | C | III.2 |
VvAAT-Jingchun1014-1 | T | A | G | A | A | G | A | C | III.2 |
VvAAT-Jingchun1014-2 | C | T | C | G | T | T | G | G | III.1 |
VvAAT-Qinlongdasui-1 | C | T | C | G | T | T | G | G | III.1 |
VvAAT-Qinlongdasui-2 | T | A | G | A | A | G | A | C | III.2 |
VvAAT-Leikejite-1 | T | A | G | G | T | T | G | G | I |
VvAAT-Leikejite-2 | T | A | G | A | A | G | A | C | II |
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