苦玄参转录组EST-SSR引物开发及群体遗传多样性分析
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摘要
目的研究18个苦玄参种群的遗传多样性及亲缘关系,为苦玄参的资源评价及开发利用提供依据。方法采用EST-SSR引物开发技术,利用SSR分子标记,分析18个种群的遗传多样性,并基于遗传距离对其进行聚类分析,明确各种群间的亲缘关系。结果从100对EST-SSR标记中,筛选出具有多态性的引物48对。随机抽取20对具有多态性的引物对18个种群进行扩增,共扩增出71个等位基因,平均每对引物3.55个。各引物间多态位点百分数(P)为0～40.7%,平均19.9%;多态信息含量(PIC)为0～0.794 1,平均0.397 7;Shannon信息指数(I)为0～1.814 3,平均0.808 4。观测杂合度(Obs_Het)为0～0.442 3,平均值0.212 7;预期杂合度(Exp_Het)变化范围0～0.826 9,平均值0.455 8。18个种群群内近交系数(Fis)值为-0.095 3～0.663 9,平均0.159 2;总群体内亚群间近交系数(Fit)值为0.062 6～0.858 7,平均0.537 2;遗传分化系数(Fst)为0～0.686,平均0.449 6。基因流(Nm)在0.114 4～0.759 4,平均值为0.306 1。各种群基因多样性指数(Nei)为0～0.401 6、I为0～0.620 9,广西梧州最高,云南普洱龙潭乡最低。云南景洪勐龙镇和云南景洪景哈乡遗传距离最近,仅0.031 9;广西龙州和云南勐海县打洛镇遗传距离最大,为0.963 8。在遗传距离0.321 3处,可将18个种群分成4个群体,其中广西3个种群归入同一群体,云南勐满镇、勐伦镇及勐遮镇归为同一个群体,第3个群体为云南勐海县勐宋乡,其余种群归入第4个群体。结论 18个种群遗传分化水平不一致,各种群杂合度差异较大。种群间Nm较小,群体基因交换不大;群体内存在一定的近交率,种群亲缘关系受地理隔离影响较大。
Objective To explore genetic diversity of and genetic relationships among 18 Picria felterrae populations to provide references for the resource assessment and utilization. Methods The genetic diversity of 18 P. felterrae populations were analyzed using the EST-SSR primer development technology and SSR molecular markers, and cluster analysis was performed based on genetic distances to determine the relationships among those populations. Results A total of 48 pairs of polymorphic primers were selected from 100 pairs of EST-SSR markers, of which 20 pairs were randomly selected and used for amplification of 18 populations. A total of 71 alleles were amplified, 3.55 alleles per primer. Among the primers, the percentage of polymorphic loci(P) varied from 0 to 40.7%, with an average of 19.9%; The polymorphism information content(PIC) varied from 0 to 0.794 1, 0.397 7 on average; The Shannon diversity information index(I) varied from 0 to 1.814 3, with an average of 0.808 4; Obs_Het varied from 0 to 0.442 3, with an average of 0.212 7; And the Exp_Het varied from 0 to 0.826 9, with an average of 0.455 8. For the 18 populations, the Inbreeding Coefficient(Fis) varied from-0.095 3 to 0.663 9, with an average of 0.159 2; The inbreeding coefficient of subgroups(Fit) varied from 0.062 6 to0.858 7, with an average of 0.537 2; The genetic differentiation coefficient(Fst) varied from 0 to 0.686, with an average of 0.449 6; The gene flow(Nm) varied from 0.114 4 to 0.759 4, with an average of 0.306 1. For the 18 samples tested, the gene diversity index(Nei) varied from 0 to 0.401 6, the I varied from 0 to 0.620 9, Wuzhou Guangxi having the maximum value and Longtan Yunnan the minimum value. Menglong and Jingha, two towns in Yunnan, had the shortest genetic distance(0.031 9), whereas Longzhou Guangxi and Menghai Yunnan had the maximum genetic distance(0.963 8). The 18 populations could be divided into four groups at the location where genetic distance was 0.321 3. The three populations in Guangxi belonged to the same group, populations from Menglong, Menglun and Mengzhe of Yunnan belonged in the same group, populations from Mengsong Yunnan became an independent group, and the rest belonged in the fourth group. Conclusion The genetic differentiation levels of 18 populations were not consistent, and the heterogeneity difference was significant. The gene flow among populations was small, which indicated that the population gene exchange was low. A certain inbreeding rate exists among the populations. The relationship among populations was influenced by geographical isolation and environmental factors.
Objective To explore genetic diversity of and genetic relationships among 18 Picria felterrae populations to provide references for the resource assessment and utilization. Methods The genetic diversity of 18 P. felterrae populations were analyzed using the EST-SSR primer development technology and SSR molecular markers, and cluster analysis was performed based on genetic distances to determine the relationships among those populations. Results A total of 48 pairs of polymorphic primers were selected from 100 pairs of EST-SSR markers, of which 20 pairs were randomly selected and used for amplification of 18 populations. A total of 71 alleles were amplified, 3.55 alleles per primer. Among the primers, the percentage of polymorphic loci(P) varied from 0 to 40.7%, with an average of 19.9%; The polymorphism information content(PIC) varied from 0 to 0.794 1, 0.397 7 on average; The Shannon diversity information index(I) varied from 0 to 1.814 3, with an average of 0.808 4; Obs_Het varied from 0 to 0.442 3, with an average of 0.212 7; And the Exp_Het varied from 0 to 0.826 9, with an average of 0.455 8. For the 18 populations, the Inbreeding Coefficient(Fis) varied from-0.095 3 to 0.663 9, with an average of 0.159 2; The inbreeding coefficient of subgroups(Fit) varied from 0.062 6 to0.858 7, with an average of 0.537 2; The genetic differentiation coefficient(Fst) varied from 0 to 0.686, with an average of 0.449 6; The gene flow(Nm) varied from 0.114 4 to 0.759 4, with an average of 0.306 1. For the 18 samples tested, the gene diversity index(Nei) varied from 0 to 0.401 6, the I varied from 0 to 0.620 9, Wuzhou Guangxi having the maximum value and Longtan Yunnan the minimum value. Menglong and Jingha, two towns in Yunnan, had the shortest genetic distance(0.031 9), whereas Longzhou Guangxi and Menghai Yunnan had the maximum genetic distance(0.963 8). The 18 populations could be divided into four groups at the location where genetic distance was 0.321 3. The three populations in Guangxi belonged to the same group, populations from Menglong, Menglun and Mengzhe of Yunnan belonged in the same group, populations from Mengsong Yunnan became an independent group, and the rest belonged in the fourth group. Conclusion The genetic differentiation levels of 18 populations were not consistent, and the heterogeneity difference was significant. The gene flow among populations was small, which indicated that the population gene exchange was low. A certain inbreeding rate exists among the populations. The relationship among populations was influenced by geographical isolation and environmental factors.
引文
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[3]王力生,马学敏,郭亚健,等.苦玄参的化学成分研究[J].中国中药杂志,2004,29(2):149-152.
[4]谢阳姣,何志鹏,闫国跃,等.苦玄参40个株系表型性状遗传多样性分析[J].广西植物,2017,37(3):348-355.
[5]谢阳姣,何志鹏,李耀燕,等.苦玄参药材中苦玄参苷IA和IB差异积累的影响因素分析[J].作物杂志,2016(3):89-93.
[6]解新明,云锦凤.植物遗传多样性及其检测方法[J].中国草地,2000(6):51-59.
[7]陈怀琼,隋春,魏建和.植物SSR引物开发策略简述[J].分子植物育种,2009,7(4):845-851.
[8]张羽.分子标记技术的产生与引物设计[J].江苏农业科学,2014,42(6):47-51.
[9]敖日格乐,贾晓,葛台明.SSR分子标记的开发策略概述[J].湖北民族学院学报:自然科学版,2009,27(4):462-467.
[10]刘果,张党权,谢耀坚,等.39种按树微卫星序列的遗传多样性分析[J].分子植物育种,2017,15(9):3767-3779.
[11]谢阳姣,李耀燕,闫国跃,等.苦玄参育种材料遗传多样性SSR分析[J].南方农业学报,2017,48(1):20-25.
[12]苏一钧,王娇,戴习彬,等.303份甘薯地方种SSR遗传多样性与群体结构分析[J].植物遗传资源学报,2018,19(2):202-210.
[13]Simon S A,Zhai J X,Nandety R S,et al.Short-read sequencing technologies transcriptional analyses[J].Annu Rev Plant Biol,2009,60:305-333.
[14]孙英新,白孝明,王丹丹.蓝莓种质资源遗传多样性的EST-SSR分析[J].辽东学院学报:自然科学版,2018,25(1):31-35.
[15]尚卫琼,段志芬,杨毅坚,等.基于EST-SSR标记的云南大叶茶资源遗传多样性分析[J].山东农业科学,2018,50(1):16-22.
[16]徐敏.中国马铃薯审定品种系谱分析及遗传多样性研究[D].北京:中国农业科学院,2007.
[17]张华丽,丛日晨,王茂良,等.基于万寿菊转录组测序的SSR标记开发[J].园艺学报,2018,45(1):159-167.
[18]陈春林,田易萍,陈林波,等.基于荧光标记的紫娟茶树转录组EST-SSR标记开发田[J].江苏农业学报,2018,34(4):747-753.
[19]逄洪波,解元坤,张璐,等.基于转录组测序的鬼针草SSR标记开发及其应用[J].分子植物育种,2018,16(16):5359-5368.