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2025年4月5日 星期六
农业生物技术学报  2022, Vol. 30 Issue (11): 2108-2118    DOI: 10.3969/j.issn.1674-7968.2022.11.005
  研究论文与报告 本期目录 | 过刊浏览 | 高级检索 |
基于RAD-seq的西番莲系统进化分析及种间SSR标记开发
田青兰1, 刘洁云1, 黄伟华1, 夏秀忠2, 杨行海2, 覃柳燕1, 张英俊1, 牟海飞1, 吴艳艳1,*
1广西壮族自治区农业科学院 生物技术研究所,南宁 530007;
2广西壮族自治区农业科学院 水稻研究所,南宁 530007
Phylogenetic Analysis and Development of SSR Markers Based on RAD-seq in Passiflora
TIAN Qing-Lan1, LIU Jie-Yun1, HUANG Wei-Hua1, XIA Xiu-Zhong2, YANG Xing-Hai2, QIN Liu-Yan1, ZHANG Ying-Jun1, MOU Hai-Fei1, WU Yan-Yan1,*
1 Biotechnology Research Institute, Guangxi Academy of Agricultural Sciences, Nanning 530007, China;
2 Rice Research Institute, Guangxi Academy of Agricultural Sciences, Nanning 530007, China
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摘要 西番莲(Passiflora spp.)是中国南方重要的果树,其种间进化关系尚不清楚,且缺乏通用性分子标记。本研究基于酶切位点相关DNA测序(restriction site-associated DNA sequencing, RAD-seq)技术对西番莲属6个种共10份种质的系统进化、SSR位点及标记通用性进行了研究。结果表明:EcoRⅠ是西番莲基因组简化测序较为适应的酶;依据剩余读长数/读长数,以'紫果7号' (P. edulis)(P4)的部分组装序列为参考基因组,利用检测到的46 451个高质量SNPs构建系统发生树,结果显示,蓝冠西番莲(P. caerulea)(P6)、红花西番莲(P. coccinea)(P7)、版纳西番莲(P. xishuangbannaensis)(P8)和大果西番莲(P. quadragularis)(P9)为一支,绿皮百香果(P. edulis)(P5)和哥伦比亚激情果(P. ligularis)(P10)为一支,金陵紫果(P. edulis)(P1)、芭乐味黄金果(P. edulis var. flavicarpa)(P2)、云南黄果原生种(P. edulis var. flavicarpa)(P3)和'紫果7号'在亲缘关系上更近;以版纳西番莲为参考基因组,利用12 452个高质量SNPs构建系统发生树,云南黄果原生种、'紫果7号'、红花西番莲、版纳西番莲、大果西番莲和哥伦比亚激情果都单独为一支,金陵紫果和芭乐味黄金果聚为一支,绿皮百香果和蓝冠西番莲聚为一支;在10份西番莲种质的基因组中共鉴定到2 614个SSR,其核心基序为AT、GA和AAG等2~6个碱基,重复数为4~16次,并成功开发了2 515对SSR引物;利用50对SSR标记评估西番莲各种质间的标记通用率为54.22%,栽培种西番莲(P1~P5)明显高于其他5个种属(P6~P10),而中国云南野生种版纳西番莲的通用率为0,推测是由于版纳西番莲与其他种质的亲缘关系较远,基因组序列上存在着较大的差异。本研究明确了不同种西番莲间的亲缘关系,为西番莲遗传改良提供理论依据,为西番莲分子标记辅助选择育种提供实用的SSR标记。
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田青兰
刘洁云
黄伟华
夏秀忠
杨行海
覃柳燕
张英俊
牟海飞
吴艳艳
关键词 西番莲SNP系统发生树SSR通用性    
Abstract:Passion fruit (Passiflora) is an important fruit tree in southern China, but the evolutionary relationship among Passiflora species is still unclear, and there is a lack of universal molecular markers to distinguish between different species. In this study, the phylogeny, SSR loci and marker versatility of 10 germplasms from 6 passion fruit species were investigated based on restriction site-associated DNA sequencing (RAD-seq). The results showed that EcoRⅠ was a suitable enzyme for simplified sequencing of passion fruit genome. According to the number of remaining reads/read lengths, the phylogenetic tree was constructed based on the 46 451 high-quality SNPs detected in RAD-seq, the partially assembled sequence of 'Purple fruit No. 7' (P. edulis)(P4) was used as the reference genome. The results showed that the blue-crowned passion fruit (P. caerulea)(P6), safflower passion fruit (P. coccinea)(P7), Banna passion fruit (P. xishuangbannaensis)(P8), and quadrangularis passion fruit (P. quadragularis)(P9) were on the same phylogenetic branch, green-skin passion fruit (P. edulis)(P5) and Colombian passion fruit (P. ligularis)(P10) were on the same branch, and Jinling purple fruit (P. edulis)(P1), guava golden fruit (P. edulis var. flavicarpa)(P2), and Yunnan yellow fruit native species (P. edulis var. flavicarpa)(P3) were closer to 'Purple fruit No. 7' in genetic relationship. When using 12 452 high-quality SNPs to construct the phylogenetic tree and using Banna passion fruit as the reference genome, the Yunnan yellow fruit native species, 'Purple fruit No. 7', safflower passion fruit, Banna passion fruit, quadrangularis passion fruit, and Colombian passion fruit were all on separate phylogenetic branches, Jinling purple fruit and guava golden fruit were on the same branch; and green-skin passion fruit and blue-crowned passion fruit were on the same branch. A total of 2 614 SSRs were identified from the 10 passion fruit germplasms, and their core motifs were AT, GA and AAG, etc., with 2~6 bases in length, and the repeat number was 4~16 times; 2 515 pairs of SSR primers were successfully developed accordingly. 50 pairs of SSR markers were used to evaluate the rate of shared markers among different passion fruit species, and the rate was 54.22%; notably, the shared rate in cultivar passion fruit (P1~P5) was significantly higher than the other 5 species (P6~P10), while the shared rate of wild Banna passion fruit in Yunnan, China was 0. This might be due to the distant genetic relationship between Banna passion fruit and other germplasms, and there were large differences in genome sequences. This study establishes the genetic relationships among different Passiflora species, provides a theoretical basis for the genetic improvement of passion fruit, and developes a useful panel of SSR markers for molecular marker-assisted selection of passion fruit.
Key wordsPassiflora    SNP    Phylogenetic tree    SSR    Transferability
收稿日期: 2021-12-09     
ZTFLH:  S323  
  S326  
基金资助:国家自然科学基金(32060660); 广西科技重大专项(桂科AA22068091-1); 广西自然科学基金(2021GXNSFBA075044; 2019GXNSFAA245002; 2018GXNSFBA281024); 广西农业科学院优势学科团队项目(桂农科2021YT089); 广西农业科学院科技发展基金(桂农科2022JM53); 广西浦北百香果试验站项目(桂TS2022010)
通讯作者: * wuyanyan@gxaas.net   
引用本文:   
田青兰, 刘洁云, 黄伟华, 夏秀忠, 杨行海, 覃柳燕, 张英俊, 牟海飞, 吴艳艳. 基于RAD-seq的西番莲系统进化分析及种间SSR标记开发[J]. 农业生物技术学报, 2022, 30(11): 2108-2118.
TIAN Qing-Lan, LIU Jie-Yun, HUANG Wei-Hua, XIA Xiu-Zhong, YANG Xing-Hai, QIN Liu-Yan, ZHANG Ying-Jun, MOU Hai-Fei, WU Yan-Yan. Phylogenetic Analysis and Development of SSR Markers Based on RAD-seq in Passiflora. 农业生物技术学报, 2022, 30(11): 2108-2118.
链接本文:  
http://journal05.magtech.org.cn/Jwk_ny/CN/10.3969/j.issn.1674-7968.2022.11.005     或     http://journal05.magtech.org.cn/Jwk_ny/CN/Y2022/V30/I11/2108
 
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