五节芒遗传多样性的ISSR分析
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摘要
五节芒(Miscanthus floridulus (Lab.)Warb.ex Schum.et Laut)为禾本科(Gramineae)芒属(Miscanthus Anderss)多年生草本植物。五节芒地下茎发达,丛状生长,生态适应性幅度极广,是一种具有开发利用价值的植物。本研究采用ISSR分子标记对五节芒12个自然居群进行了遗传多样性和遗传结构的分析,目的是为了解五节芒居群的遗传多样性水平、遗传结构以及居群间的遗传分化,为五节芒资源的保护与开发利用提供理论依据。主要结果如下：
建立了适合于五节芒的ISSR-PCR反应体系和反应程序。25μL体系中,10×PCR Buffer 2.5μL、Taq DNA聚合酶0.75 U.Mg2+1.5mmol/L、4×dNTP 0.2 mmol/L。PCR反应程序为：94℃预变性5 min；94℃变性45 s；50-60℃退火45s,72℃延伸1 min 20 s,45个循环；72℃再延伸7 min；4℃恒温保存。
利用7条ISSR引物对五节芒12个居群共186个个体进行了扩增,共得到88条谱带,其中多态性条带为81条,结果表明,五节芒在物种水平的遗传多样性较高(PPF=92.05%;Na=1.9205,Ne=1.4941,Hpop=0.2911,I=0.4409)。分析表明,居群水平上的遗传多样性相对较低(PPF=43.85%,Na= 1.4385,Ne=1.2564,Hpop=0.1534,I=0.2286),其中靖安居群的遗传多样性最高。
五节芒居群间存在较大的遗传分化,居群间的遗传分化系数Gst=0.4669,表明遗传变异主要表现在居群内。五节芒为多年生植物,分布范围广；生殖方式为有性生殖和营养繁殖,这些可能导致较高遗传多样性水平和较低居群遗传分化。
UPGMA聚类分析和POPGEN结果把五节芒分成三支。经Mantel检验遗传距离与地理距离没有显著相关性(r=0.3086)。
Miscanthus floridulus (Lab.) Warb. ex Schum. et Laut is an economically important plant. The study were to evaluate the genetic diversity and genetic differentiation of natural populations and present a scientific framework for the conservation and utilization of M. floridulus.
We established the reaction system and amplified procedure suitable for M. floridulus, which was 25μL amplification reactions system containing 10×PCR buffer 2.5μL,1.5 mmol/L Mg2+,100 ng template DNA,200 mmol/L dNTPs,0.75 U Taq DNA polymerase. The optimal amplified procedure was as follows:after a pre-denaturing of 5 min at 94℃,45 cycles were performed with denaturing of 45 s at 94℃, annealing of 45 s due from 50-60℃, extension of 80 s at 72℃, a final extension step of 7 min at 72℃.
We estimated the genetic diversity and genetic structure of 12 natural populations in Jiangxi province using ISSR. A relatively high level of genetic diversity was detected in M. floridulus using 81 polymorphic microsatellite locie. The ISSR analyses revealed a high level of genetic diversity at the species level (PPF= 92.05%,Na=1.9205, Ne=1.4941, Hpop=0.2911,I=0.4409); and a low level at the populations level (PPF=43.85%,Na=1.4385, Ne=1.2564, Hpop=0.1534, I=0.2286). The genetic diversity of Jing'an population(JA) was the highest.
M. floridulus is a perennial species with a wide distribution; these characteristics could account for the relatively high genetic diversity we observed. Furthermore this species has outcrossing and incrossing breeding system, a situation amenable to high genetic diversity and low levels of genetic differen-tiation among populations (Gst= 0.4669). Divided into 3 groups, using UPGMA and POPGEN. Mantel test evealed no significant correlation between the genetic distance and geographic distance (r= 0.3086).
建立了适合于五节芒的ISSR-PCR反应体系和反应程序。25μL体系中,10×PCR Buffer 2.5μL、Taq DNA聚合酶0.75 U.Mg2+1.5mmol/L、4×dNTP 0.2 mmol/L。PCR反应程序为：94℃预变性5 min；94℃变性45 s；50-60℃退火45s,72℃延伸1 min 20 s,45个循环；72℃再延伸7 min；4℃恒温保存。
利用7条ISSR引物对五节芒12个居群共186个个体进行了扩增,共得到88条谱带,其中多态性条带为81条,结果表明,五节芒在物种水平的遗传多样性较高(PPF=92.05%;Na=1.9205,Ne=1.4941,Hpop=0.2911,I=0.4409)。分析表明,居群水平上的遗传多样性相对较低(PPF=43.85%,Na= 1.4385,Ne=1.2564,Hpop=0.1534,I=0.2286),其中靖安居群的遗传多样性最高。
五节芒居群间存在较大的遗传分化,居群间的遗传分化系数Gst=0.4669,表明遗传变异主要表现在居群内。五节芒为多年生植物,分布范围广；生殖方式为有性生殖和营养繁殖,这些可能导致较高遗传多样性水平和较低居群遗传分化。
UPGMA聚类分析和POPGEN结果把五节芒分成三支。经Mantel检验遗传距离与地理距离没有显著相关性(r=0.3086)。
Miscanthus floridulus (Lab.) Warb. ex Schum. et Laut is an economically important plant. The study were to evaluate the genetic diversity and genetic differentiation of natural populations and present a scientific framework for the conservation and utilization of M. floridulus.
We established the reaction system and amplified procedure suitable for M. floridulus, which was 25μL amplification reactions system containing 10×PCR buffer 2.5μL,1.5 mmol/L Mg2+,100 ng template DNA,200 mmol/L dNTPs,0.75 U Taq DNA polymerase. The optimal amplified procedure was as follows:after a pre-denaturing of 5 min at 94℃,45 cycles were performed with denaturing of 45 s at 94℃, annealing of 45 s due from 50-60℃, extension of 80 s at 72℃, a final extension step of 7 min at 72℃.
We estimated the genetic diversity and genetic structure of 12 natural populations in Jiangxi province using ISSR. A relatively high level of genetic diversity was detected in M. floridulus using 81 polymorphic microsatellite locie. The ISSR analyses revealed a high level of genetic diversity at the species level (PPF= 92.05%,Na=1.9205, Ne=1.4941, Hpop=0.2911,I=0.4409); and a low level at the populations level (PPF=43.85%,Na=1.4385, Ne=1.2564, Hpop=0.1534, I=0.2286). The genetic diversity of Jing'an population(JA) was the highest.
M. floridulus is a perennial species with a wide distribution; these characteristics could account for the relatively high genetic diversity we observed. Furthermore this species has outcrossing and incrossing breeding system, a situation amenable to high genetic diversity and low levels of genetic differen-tiation among populations (Gst= 0.4669). Divided into 3 groups, using UPGMA and POPGEN. Mantel test evealed no significant correlation between the genetic distance and geographic distance (r= 0.3086).
引文
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