甜瓜抗病基因同源序列的克隆与分析Cloning and Analysis of Melon Resistance Gene Analogs
王贤磊,高兴旺,张铁钢,王文林,熊丽曼,韩瑞,李冠
摘要(Abstract)：
为了获得新的甜瓜抗病基因同源基因(RGA),我们根据NBS-LRR类抗病基因保守区设计兼并引物组合,以抗病甜瓜品种基因组DNA为模板进行PCR扩增,经筛选、分类与基因测序,获得7个甜瓜RGA序列,其中821与324两个RGA为本研究首次发现.将得到的序列与已知甜瓜RGA序列进行分析、比对,将重复的RGA合并,通过对不同RGA序列同源分析,发现甜瓜CC-NBS-LRR类与TIR-NBS-LRR类RGA的不同分布特征与不同RGA之间的相互关系,如MRGA10与MRGH5可能起源于同一基因.通过对RGA的遗传分析与比较基因组学分析,发现甜瓜RGA序列与抗病基因之间的对应关系,如抗枯萎病基因Fom-1和抗番木瓜环斑病基因Prv存在于nbs47-3(MRGH11)与MRGH21之间,其中候选抗病基因MRGH8推测为Fom-1;候选抗病基因MRGH9、MRGH10之一为Prv.
关键词(KeyWords)： 抗病基因;甜瓜;同源序列;克隆
基金项目(Foundation): 国家自然科学基金(30660078,31060148);; 新疆维吾尔自治区高技术项目(200711102)资助
作者(Author): 王贤磊,高兴旺,张铁钢,王文林,熊丽曼,韩瑞,李冠
参考文献(References)：
- [1]Young ND.The genetic architecture of resistance[J].Curr Opin Plant Biol,2000,3(4):285-290.
- [2]Meyers BC,Kozik A,Griego A,et al.Genome-Wide Analysis of NBS-LRR Encoding Genes in Arabidopsis[J].The PlantCell,2003,15:809–834.
- [3]Bai J,Pennill LA,Ning J,et al.Diversity in nucleotide binding site-leucine-rich repeat genes in cereals[J].Genome Res,2002,12:1871-1884.
- [4]Hulbert SH,Webb CA,Smith SM,et al.Resistance gene complexes:evolution and utilization[J].Annu Rev Phytopathol,2001,39:285-312.
- [5]Garcia-Mas J,Leeuwen H,Monfort A,et al.Cloning and mapping of resistance gene homologues in melon[J].Plant Science,2001,161(1):165-172.
- [6]Leeuwen V,Garcia-Mas J,Coca M,et al.Analysis of the melon genome in regions encompassing TIR-NBS-LRR resistancegenes[J].Mol Genet Genomics,2005,273(3):240-251.
- [7]Brotman Y,Silberstein L,Kovalski I,et al.Resistance gene homologues in melon are linked to genetic loci conferringdisease and pest resistance.[J]Theor Appl Genet,2002,104(6-7):1055-1063.
- [8]Tezuka T,Waki K,Yashiro K,et al.Construction of a linkage map and identification of DNA markers linked to Fom-1,agene conferring resistance to Fusarium oxysporum f.sp melonis race 2 in melon[J].Euphytica,2009,168(2):177-188.
- [9]Daryono BS,Development and application of melon resistance gene analogs(MRGA)marker for detection of virus resistancegene(creb-2)in melon[C].The 4th international cucurbitaceae symposium,2009,20-24.
- [10]Joobeur T,King JJ,Nolin SJ,et al.The fusarium wilt resistance locus Fom-2 of melon contains a single resistance genewith complex features[J].Plant Journal,2004,39(3):283-297.
- [11]Pauquet J,Burget E,Hagen L.Map-based cloning of the Vat gene from melon conferring resistance to both aphid coloniza-tion and aphid transmission of several viruses[C].Proc Cucurbitaceae 2004,8th Eucarpia Meeting on Cucurbit Geneticsand Breeding,Olomouc,2004,325-329.
- [12]Sue P,Grant B,Bernard RB,Modification of a CTAB DNA extraction protocol for plants containing high polysaccharideand polyphenol components[J].Plant Mol Biol Rep,1997,15(1):8-15.
- [13]萨姆布鲁克,拉塞尔.分子克隆实验指南[M].黄培堂译.第3版,北京:科学出版社,2002,96-99.
- [14]Huang SW,Li RQ,Zhang ZH et al.The genome of the cucumber[J],Cucumis sativus L.Nat Genet,2009,41(12):1275-1283.
- [15]Brotman Y,Kovalski,Dogimont,et al.Molecular markers linked to papaya ring spot virus resistance and Fusarium race 2resistance in melon[J].Theor Appl Genet,2005,110(2):337-345.
- [16]Wang Xianlei,Li Guan,Gao Xingwang,et al.Powdery mildew resistance gene(Pm-AN)located in a segregation distortionregion of melon LGV[J].Euphytica,2011,published on line.DOI:10.1007/s10681-011-0406-1