基于SRAP分子标记的天麻遗传多样性研究
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摘要
目的利用SRAP分子标记技术对天麻种质资源进行遗传多样性研究,为天麻不同亲缘物种间的分类及其优良种质资源的筛选提供依据。方法采集7个不同生态区域的天麻种质资源,包括红杆G.elata f.elata、乌杆G.elata f.glauca和绿杆G.elata f.viridis 3种变型和红乌杂交天麻,共24份样品。运用SRAP分子标记方法构建天麻种质的DNA指纹图谱,从分子水平检测其遗传多样性。结果 33对引物扩增出637条多态性条带,多态性百分率达73.16%。24份天麻种质样品间相似度分布于0.404 0~0.908 0,整个群体之间的相似程度差异较大。其中红天麻样品间的相似度在0.906 6~0.996 4,遗传差异较小;乌天麻、杂交天麻和绿天麻样品间的相似度分别在0.410 4~0.999 6、0.541 0~0.950 4和0.578 2,遗传差异较大。AMOVA分析结果显示,天麻变型内变异大于变型间变异,天麻各变型间有很大的遗传分化(FST=0.33,P<0.05)。另外,人工栽培对遗传分化有影响,但不显著。结论 SRAP分子标记方法得到的24份天麻样品多态性丰富,能有效地反映出天麻的遗传多样性。红杆天麻的遗传性状较为稳定,与其他变型间缺乏基因交流,遗传多样性匮乏;其他2个变型具有较高的遗传多样性。
Objective In order to correctly identify the different germplasm resources of Gastrodia elata and gain the excellent germplasm resources, SRAP molecular marker was used to analyze the genetic diversity of G. elata. Methods G. elata was collected from seven different areas, which included 24 smaples of G. elata f. elata, G. elata f. g1 auca and G. elata f. viridis, the DNA figerprint of G. elata was constructed with SRAP molecular marker, and the genetic diversity was analyzed. Results Six huandred Thirty-seven belts were amplified by 33 primers pairs, and the polymorphic percentage was 73.16%. The range of genetic similarity coefficient was 0.404 0—0.908 0, the genetic similarity coefficient among G. elata f. elata was in the range of 0.906 6—0.996 4, and those of G. elata f. g1 auca, G. elata f. viridis, and hybrid was in the range of 0.410 4—0.999 6, 0.541 0—0.950 4 and 0.578 2, respectively. They showed small genetic differences. The analysis of molecular variance showed higher percentages of genetic variation within population than those among populations in all species. Populations showed higher genetic structure(FST = 0.33, P < 0.05). In addition, artificial cultivation had influence to the genetic differentiation, but not significantly. So in terms of different cultivation conditions so far had no significant impact on the genetic differentiation of G. elata. Mantel's test has been used to detect the correlation between genetic distance and geographic distance of all sorts of germplasm resources, the result had no significant correlation, and due to the limited number of samples, the result is not representative. Conclusion SRAP molecular marker method can more effectively reflect the genetic polymorphisms of G. elata. Compared with other two phenotypes, G. elata f. elata is more conservative and has lower genetic diversity. The other two variants have higher genetic diversity.
Objective In order to correctly identify the different germplasm resources of Gastrodia elata and gain the excellent germplasm resources, SRAP molecular marker was used to analyze the genetic diversity of G. elata. Methods G. elata was collected from seven different areas, which included 24 smaples of G. elata f. elata, G. elata f. g1 auca and G. elata f. viridis, the DNA figerprint of G. elata was constructed with SRAP molecular marker, and the genetic diversity was analyzed. Results Six huandred Thirty-seven belts were amplified by 33 primers pairs, and the polymorphic percentage was 73.16%. The range of genetic similarity coefficient was 0.404 0—0.908 0, the genetic similarity coefficient among G. elata f. elata was in the range of 0.906 6—0.996 4, and those of G. elata f. g1 auca, G. elata f. viridis, and hybrid was in the range of 0.410 4—0.999 6, 0.541 0—0.950 4 and 0.578 2, respectively. They showed small genetic differences. The analysis of molecular variance showed higher percentages of genetic variation within population than those among populations in all species. Populations showed higher genetic structure(FST = 0.33, P < 0.05). In addition, artificial cultivation had influence to the genetic differentiation, but not significantly. So in terms of different cultivation conditions so far had no significant impact on the genetic differentiation of G. elata. Mantel's test has been used to detect the correlation between genetic distance and geographic distance of all sorts of germplasm resources, the result had no significant correlation, and due to the limited number of samples, the result is not representative. Conclusion SRAP molecular marker method can more effectively reflect the genetic polymorphisms of G. elata. Compared with other two phenotypes, G. elata f. elata is more conservative and has lower genetic diversity. The other two variants have higher genetic diversity.
引文
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[15]王世峰,王军.斜带髭鲷野生与养殖群体遗传结构的ISSR分析[J].海洋学报,2007,29(4):105-110.
[16]段艳凤,金黎平.中国88个马铃薯审定品种SSR指纹图谱构建与遗传多样性分析[J].作物学报,2009,35(8):1451-1457.
[17]Aneja B,Neelam R,Chawla V,et al.Sequence-related amplified polymorphism(SRAP)molecular marker system and its applications in crop improvement[J].Mol Breed,2012,30:1635-1648.
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[19]Mutlu N,Abak K.Development of SRAP,SRAP-RGA,RAPD and SCAR markers linked with a Fusarium wilt resistance gene in eggplant[J].Theor Appl Genet,2008,117:1303-1312.
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[21]Ku C,Chung W C.The Complete Plastid genome sequence of madagascar Periwinkle catharanthus roseus(L.)G.Don:plastid genome evolution,molecular marker identification,and phylogenetic implications in asterids[J].PLoS One,2013,8(6):e6858.
[22]尹海波,王吉华,涂秀文,等.不同产地老鹳草遗传多样性ISSR分析[J].中草药,2013,44(22):3206-3211.
[23]阳翠,刘萍,刘姣蓉,等.苦豆子ISSR标记的遗传多样性分析[J].中草药,2013,44(10):1323-1327.
[24]关萍,石建明.不同分布区天麻的AFLP分析[J].植物遗传资源学报,2013,14(1):66-73.
[25]谢渊,张小蕾.AFLP技术在天麻遗传变异研究中的初步应用[J].植物生理学通讯,2007,43(1):141-144.
[26]陈祖云,王晓丽.贵州天麻遗传多态性的ISSR初步分析[J].中华中医药杂志,2007,22(7):436-439.
[27]王德信.天麻ITS序列分析及变异类型鉴定[J].生物技术,2010,20(6):33-35.
[28]李作洲,暴朝霞,黄宏文.药用植物天麻(Gastrodia elata Bl.)的等位酶遗传变异及其变型间的遗传关系[J].植物科学学报,2011,29(1):64-73.
[29]王德信.天麻ITS-1测序及单核苷酸多态性变异位点分析[J].生物技术,2012,22(5):48-51.
[30]徐锦堂.家栽天麻与野生天麻质量比较[J].中国药学杂志,2000,35(8):511-513.
[2]周铉.当今药市上的各种天麻[M].北京:中国科学院药用植物研究所,2013.
[3]徐锦堂.中国天麻栽培学[M].北京:北京医科大学中国协和医科大学联合出版社,1993.
[4]房超,李跃建.应用SRAP分子标记构建茄子遗传图谱初探[J].西南农业学报,2010,28(5):1591-1594.
[5]杨世林,兰进,徐锦堂.天麻的研究进展[J].中草药,2000,31(1):66-69.
[6]刘小琴,汪鋆植,袁琴.天麻不同品种及不同组织中天麻素的含量比较[J].时珍国医国药,2009,20(4):908-909.
[7]刘明学,李琼芳,李梁.平武天麻GAP基地三种天麻变异类型质量评价[J].现代中药研究与实践,2009,23(5):3-5.
[8]李德勋,陈桂,等.天麻不同变异类型药材中天麻素含量比较[J].现代中药研究与实践,2007,22(3):23-24.
[9]Chen W,Zhang Y.Detection of QTL for six yield-related traits in oilseed rape(Brassica napus)using DH and immortalized F2 populations[J].Theor Appl Genet,2007,115:849-858.
[10]Lu Y Q,Wu W R.Identification of salt-responsive genes in English cordgrass(Spartina anglica)roots using SRAP technique[J].J Zhejiang Univ,2006,32(5):511-514.
[11]栾丽.同源四倍体水稻三系亲本的遗传差异、细胞遗传学比较研究及低直链淀粉含量突变体Wx基因序列分析[D].成都:中科院研究生院,2007.
[12]邓传良,曹莹.菠菜性别相关SRAP分子标记的筛选及分析[J].基因组学与应用生物学,2013,32(4):516-521.
[13]胡秀,刘念.中国姜花属基于SRAP分子标记的聚类分析[J].植物分类学报,2008,46(6):899-905.
[14]刘雅辉,刘大群.23个小麦抗叶锈近等基因系SRAP多态性[J].中国农业科学,2008,41(5):1333-1340.
[15]王世峰,王军.斜带髭鲷野生与养殖群体遗传结构的ISSR分析[J].海洋学报,2007,29(4):105-110.
[16]段艳凤,金黎平.中国88个马铃薯审定品种SSR指纹图谱构建与遗传多样性分析[J].作物学报,2009,35(8):1451-1457.
[17]Aneja B,Neelam R,Chawla V,et al.Sequence-related amplified polymorphism(SRAP)molecular marker system and its applications in crop improvement[J].Mol Breed,2012,30:1635-1648.
[18]陈名红,李玉.分子标记技术在药用植物研究中的应用前景[J].贵州农业科学,2011,39(2):19-22.
[19]Mutlu N,Abak K.Development of SRAP,SRAP-RGA,RAPD and SCAR markers linked with a Fusarium wilt resistance gene in eggplant[J].Theor Appl Genet,2008,117:1303-1312.
[20]Li G,Quiros C F.Sequence-related amplified polymor-phism(SRAP),a new marker system based on a simple PCR reaction:its application to mapping and gene tagging in Brassica[J].Theor Appl Genetics,2001,103:455-461.
[21]Ku C,Chung W C.The Complete Plastid genome sequence of madagascar Periwinkle catharanthus roseus(L.)G.Don:plastid genome evolution,molecular marker identification,and phylogenetic implications in asterids[J].PLoS One,2013,8(6):e6858.
[22]尹海波,王吉华,涂秀文,等.不同产地老鹳草遗传多样性ISSR分析[J].中草药,2013,44(22):3206-3211.
[23]阳翠,刘萍,刘姣蓉,等.苦豆子ISSR标记的遗传多样性分析[J].中草药,2013,44(10):1323-1327.
[24]关萍,石建明.不同分布区天麻的AFLP分析[J].植物遗传资源学报,2013,14(1):66-73.
[25]谢渊,张小蕾.AFLP技术在天麻遗传变异研究中的初步应用[J].植物生理学通讯,2007,43(1):141-144.
[26]陈祖云,王晓丽.贵州天麻遗传多态性的ISSR初步分析[J].中华中医药杂志,2007,22(7):436-439.
[27]王德信.天麻ITS序列分析及变异类型鉴定[J].生物技术,2010,20(6):33-35.
[28]李作洲,暴朝霞,黄宏文.药用植物天麻(Gastrodia elata Bl.)的等位酶遗传变异及其变型间的遗传关系[J].植物科学学报,2011,29(1):64-73.
[29]王德信.天麻ITS-1测序及单核苷酸多态性变异位点分析[J].生物技术,2012,22(5):48-51.
[30]徐锦堂.家栽天麻与野生天麻质量比较[J].中国药学杂志,2000,35(8):511-513.