中国食用向日葵种质资源遗传变异的RAPD及AFLP分析

本研究采用RAPD和AFLP方法对23个中国不同地区的食用向日葵(Helianthus annuus L.)骨干品种进行了遗传变异分析,同时对两种标记系统进行了比较.26个RAPD引物产生了总计192条DNA条带,大小分布于0.26 kb～1.98 kb之间,其中165条(86.12%)具有多态性,每条引物产生DNA条带的平均数为7.38.8对AFLP引物组合共产生了576条带,分布于100 bp～500 bp之间,其中的341条具有多态性,多态百分率为76.00%,每对引物组合产生DNA条带的平均数为72.RAPD方法检测到的每位点有效等位基因数(1.76)大于AFLP(1.65),AFLP标记位点的平均多态性信息量(PIC)(0.38)低于RAPD标记位点的PIC(0.41),但AFLP标记具有很高的多态性检测效率(Ai=38.52).用RAPD标记分析23个食用向日葵材料的亲缘关系,Nei氏相似性系数分布在47.84%～82.06%,平均相似性系数为0.649 5, 而采用AFLP的Nei氏相似性系数分布在54.15%～83.52%,平均相似性系数为0.688 4.RAPD数据的标准差为0.13,而AFLP数据的标准差为0.08.因此,采用RAPD和AFLP方法分析食用向日葵遗传变异,RAPD标记具有较低相似性系数和较高方差而AFLP则相反.源于两种不同标记的遗传相似性矩阵的相关系数为0.51,说明采用RAPD和AFLP系统分析食用向日葵遗传变异得到的结果有一定的相关性.无论采用RAPD还是AFLP标记进行聚类分析,都将23个不同基因型的食用向日葵材料分成了三个类群.     

名贵茶花种质资源的RAPD分析

采用RAPD方法构建了国内外23个茶花品种的指纹图谱。从100个随机引物中筛选出的20个有效引物共产生136条DNA片段,遗传多态性带120条占总数88．2％。遗传相似性分析表明,各基因型间的Nei＇s相似系数分布在0．4386～0．8936之间,平均相似性系数为0．7668。通过非加权算术平均数聚类（UPGMA）法,绘制了它们的遗传关系树状图,23个品种可划分为3个类群。研究结果表明,RAPD技术可用于茶花品种的鉴别以及种质资源遗传多态性的研究。     

羊草种质基因组DNA的AFLP多态性研究

羊草是禾本科牧草之王 ,在当前我国西部生态建设和草原畜牧业发展中发挥着重要作用。用AFLP方法对2 7份我国不同地区分布的羊草 (Leymuschinensis (Trin .)Tzvel)材料进行了基因组DNA多态性分析 ,8对AFLP引物组合在 2 7个不同羊草基因型中共扩增出 5 37条带 ,产生出的DNA片段大小分布在 75bp - 5 30bp之间。其中单态性带 89条 ,占 16 .6 % ,多态性带 32 9条 ,占 6 1.3%。平均每对引物组合扩增的DNA带数为 6 6 .13,总的多态性比率为 78.84%。AFLP多态信息含量PIC值分布于 0 .0 - 0 .5之间 ,平均PIC值为 0 .2 16 ,出现的PIC最大值 (0 .5 )约占AFLP标记的 8.5 % ,说明羊草基因组DNA的多态性比较丰富。以 5 37个AFLP标记为原始数据 ,根据Nei和Li的方法对 2 7份羊草材料进行遗传变异和聚类分析的结果表明 :羊草种内有高频率的遗传变异发生 ,且与地理分布和生态环境密切相关 ;2 7份羊草不同基因型被划分为四大类群 ,不同类群相互间的遗传距离相对较大 ,在树状图中表现为较远的亲缘关系。对羊草种内遗传变异发生的原因和品种的形成进行了初步讨论。     

羊草种质基因组DNA的AFLP多态性研究

羊草是禾本科牧草之王,在当前我国西部生态建设和草原畜牧业发展中发挥着重要作用.用AFLP方法对27份我国不同地区分布的羊草(Leymus chinensis (Trin.) Tzvel)材料进行了基因组DNA多态性分析,8对AFLP引物组合在27个不同羊草基因型中共扩增出537条带,产生出的DNA片段大小分布在75 bp-530 bp之间.其中单态性带89条,占16.6%,多态性带329条,占61.3%.平均每对引物组合扩增的DNA带数为66.13,总的多态性比率为78.84%.AFLP多态信息含量PIC值分布于0.0-0.5之间,平均PIC值为0.216,出现的PIC最大值(0.5)约占AFLP标记的8.5%,说明羊草基因组DNA的多态性比较丰富.以537个AFLP标记为原始数据,根据Nei和Li的方法对27份羊草材料进行遗传变异和聚类分析的结果表明:羊草种内有高频率的遗传变异发生,且与地理分布和生态环境密切相关;27份羊草不同基因型被划分为四大类群,不同类群相互间的遗传距离相对较大,在树状图中表现为较远的亲缘关系.对羊草种内遗传变异发生的原因和品种的形成进行了初步讨论.     

附子野生资源群体遗传多样性的RAPD分析

应用RAPD标记分析了分布在附子主栽区四川、重庆、陕西及湖北16个野生乌头种群的遗传变异。24个引物共检测到643个RAPD位点,多态位点602个,总的多态位点百分率达93．5％。Shannon多样性和Nei遗传分化结果一致显示重庆酉阳种群和重庆城口种群遗传多样性最高,四川盐源种群和陕西勉县种群的遗传多样性最低。Shannon指数测出的种群内的遗传变异（57．6％）略占优势,群体间的遗传分化达到42．4％;Nei基因分化系数（GST）达40．0％;分子方差分析（AMOVA）发现群体间遗传变异仅为25．37％;种群每代迁移数Nm为0．756。Nei相似性系数的UPGMA分析结果显示该地区的野生乌头分布上有一定的地域性,特别是同为附子道地产地江油供种的北川、安县和青川种群间遗传关系密切,说明种质资源在道地药材形成中具有重要作用。研究结果表明,附子主要栽培地区的乌头野生种群之间存在较大的遗传分化,遗传多样性较高,遗传种质资源较丰富,存在一定的特异性资源,为进一步开发利用乌头（川乌、附子）提供了丰富的种质资源。     

AFLP和RAPD标记技术在栉孔扇贝遗传多样性研究中的应用比较

AFLP和RAPD标记技术是近年来发展最快的基于PCR基础上的两种DNA标记技术,本文比较了两种标记技术在我国栉孔扇贝群体遗传多样性研究中的应用。共筛选20个RAPD引物和7个AFLP引物组合,检测到AFLP标记的有效等位基因数和平均多态信息量稍低于RAPD标记,但AFLP标记在每单位分析中扩增到的野生和养殖群体的多态性条带数(23．8,24．8)分别高于RAPD标记(5．6,5．6),AFLP多态性检测效率显著高于RAPD标记。AFLP和RAPD两种标记技术所揭示的野生种群与养殖群体间的近交系数、遗传距离两项指标均表明,我国栉孔扇贝养殖群体和野生种群之间尚未出现明显的遗传分化。研究结果表明：RAPD和AFLP这两种标记技术均可用于栉孔扇贝遗传多样性的分析,其分析结果是一致的。     

黄淮麦区以1B/1R类品种为抗源主要育成小麦品种的遗传多样性分析

采用SSR技术对黄淮麦区以1B／1R类品种为抗源育成的38个小麦品种进行聚类分析。39个SSR引物共扩增出186条谱带,其中143务为多态性条带,占76．9％。每个引物可扩增出1～9条多态性条带,平均3．7条。位点多态性信息含量PIC变幅为0．320-0．857,平均为0．634。聚类分析表明,在遗传距离GD值0．32水平上38个小麦品种可聚成六大类。品种间遗传距离GD变幅为0．10769～0．48571。SSR标记揭示出这38个具有黑麦血缘的小麦品种遗传变异较小,遗传基础比较狭窄。     

河北省大豆推广品种遗传多样性分析

利用主要农艺性状以及SSR和AFLP2种分子标记,对河北省41个大豆推广品种进行遗传多样性分析,以便为种质资源利用和创新提供依据。农艺性状聚类结果将41个材料划分为3个类群和2个特殊品种,聚类结果与材料系谱来源相差悬殊,不能反映材料间亲缘关系。SSR和AFLP数据聚类结果将41个材料划分为4个SAG（SSR and AFLP—basedgroups）分子类群。30对SSR引物共检测出135个等位变异,平均每个位点上有4．47个等位变异,SSR的遗传多样性指数（Simpson）分布范围为0．0928～0．7800,平均值为0、6442。10对AFLP引物共扩增出93个多态性标记,平均每对引物9．3个多态性标记。品种间的遗传相似系数（GS）变化范围为0．5877～0．9868,平均值变化范围为0．6732～0．7653,总体平均值为0.7237,遗传相似系数较高,说明材料间遗传变异较小。     

西南地区野生狗牙根种质资源的SSR与AFLP联合分析

为了制定科学的资源保护策略、促进其开发利用以及新品种的选育,对西南地区55份野生狗牙根材料,联合采用 SSR 和 AFLP 方法分析其遗传多样性.通过进行引物筛选,筛选出有效引物组合18对和11对;SSR 和 AFLP 扩增总条带353和626条,多态性条带267和592条,平均每对引物扩增出多态性条带19．61和53．83条,多态性位点百分率为75．10％和94．57％.聚类分析表明 GS＝0．806时,可将所有供试材料分成6个组群,当 GS＝0．82时,可将第 VI 个组群分成4个小组,当 GS＝0．836时,第 VI 组群中第3小组的14个材料分为3个组群基于 Shannon 多样性指数,类群内的遗传变异占总变异的67．41％,类群间的遗传变异占总变异的32．59％.基于 Nei 氏无偏估计的遗传一致度的聚类分析表明,各生态地理类群间的遗传分化与其所处的生态地理环境具有一定的相关性.通过两种标记的相关性分析,表明 SSR 和 AFLP 标记之间具有显著的相关性,而且相关性较高,证明供试材料的聚类和其生态地理环境间有一定的相关性;采用 SSR 和 AFLP 两种联合分析,结果会更加合理和有效.     

木霉肽类代谢产物对豇豆土著根瘤菌遗传性状的影响

利用RAPD与ISSR分子标记检测手段,分析了哈茨木霉T2-16肽类代谢产物处理豇豆土著根瘤菌,对其遗传性状的影响,同时,比较了RAPD和ISSR两种不同分子标记在检测根瘤菌种间的遗传相似性以及遗传变异性的分辨力.实验中,从100条引物中筛选到具有多态性的ISSR引物5条,从80条引物中筛选到具有多态性的RAPD引物6条,用5条ISSR引物扩增出54条带,多态性条带比率为75.93 %;6条RAPD引物扩增出61条带,多态性条带比率为68.85 %.两种分子标记均能揭示出处理前后根瘤菌间的遗传差异,但ISSR标记比RAPD标记可检测到更大的遗传变异.根据两种标记的结果,对供试的根瘤菌进行聚类分析,结果表明,土著根瘤菌经木霉肽类代谢产物处理后,与出发菌株相比,表现出一定程度的遗传分化和遗传差异性.     

Comparative analysis on the genetic relatedness of Sorghum bicolor accessions from Southern Africa by RAPDs,AFLPs and SSRs

In order to get an overview on the genetic relatedness of sorghum (Sorghum bicolor) landraces and cultivars grown in low-input conditions of small-scale farming systems, 46 sorghum accessions derived from Southern Africa were evaluated on the basis of amplified fragment length polymorphism (AFLPs), random amplified polymorphic DNAs (RAPDs) and simple sequence repeats (SSRs). By this approach all sorghum accessions were uniquely fingerprinted by all marker systems. Mean genetic similarity was estimated at 0.88 based on RAPDs, 0.85 using AFLPs and 0.31 based on SSRs. In addition to this, genetic distance based on SSR data was estimated at 57 according to a stepwise mutation model (Deltamu-SSR). All UPGMA-clusters showed a good fit to the similarity estimates (AFLPs: r = 0.92; RAPDs: r = 0.88; SSRs: r = 0.87; Deltamu-SSRs: r = 0.85). By UPGMA-clustering two main clusters were built on all marker systems comprising landraces on the one hand and newly developed varieties on the other hand. Further sub-groupings were not unequivocal. Genetic diversity (H, DI) was estimated on a similar level within landraces and breeding varieties. Comparing the three approaches to each other, RAPD and AFLP similarity indices were highly correlated (r = 0.81), while the Spearman's rank correlation coefficient between SSRs and AFLPs was r = 0.57 and r = 0.51 between RAPDs and SSRs. Applying a stepwise mutation model on the SSR data resulted in an intermediate correlation coefficient between Deltamu-SSRs and AFLPs (r = 0.66) and RAPDs ( r = 0.67), respectively, while SSRs and Deltamu-SSRs showed a lower correlation coefficient (r = 0.52). The highest bootstrap probabilities were found using AFLPs (56% on average) while SSR, Deltamu-SSR and RAPD-based similarity estimates had low mean bootstrap probabilities (24%, 27%, 30%, respectively). The coefficient of variation (CV) of the estimated genetic similarity decreased with an increasing number of bands and was lowest using AFLPs.     

Assessment of genetic variation withinBrassica campestris cultivars using amplified fragment length polymorphism and random amplification of polymorphic DNA markers

Genetic relationships were evaluated among nine cultivars ofBrassica campestris by employing random amplification of polymorphic DNA (RAPD) and amplified fragment length polymorphism (AFLP) markers. RAPDs generated a total of 125 bands using 13 decamer primers (an average of 9.6 bands per assay) of which nearly 80% were polymorphic. The per cent polymorphism ranged from 60–100%. AFLP, on the other hand generated a total of 319 markers, an average of 64 bands per assay. Of these, 213 were polymorphic in nature (66.8%). AFLP methodology detected polymorphism more efficiently than RAPD approach due to a greater number of loci assayed per reaction. Cultivar-specific bands were identified, for some cultivars using RAPD, and for most cultivars with AFLP. Genetic similarity matrix, based on Jaccard’s index detected coefficients ranging from 0.42 to 0.73 for RAPD, and from 0.48 to 0.925 for AFLPs indicating a wide genetic base. Cluster analyses using data generated by both RAPD and AFLP markers, clearly separated the yellow seeded, self-compatible cultivars from the brown seeded, self-incompatible cultivars although AFLP markers were able to group the cultivars more accurately. The higher genetic variation detected by AFLP in comparison to RAPD was also reflected in the topography of the phenetic dendrograms obtained. These results have been discussed in light of other studies and the relative efficiency of the marker systems for germplasm evaluation.     

利用RFLP、SSR、AFLP和RAPD标记分析玉米自交系遗传多样性的比较研究

利用 ＲＦＬＰ、ＳＳＲ．ＡＦＬＰ和ＲＡＰＤ ４种分子标记方法研究了 １５个玉米（Ｚｅａ ｍａｙｓ Ｌ．）自交系的遗传多样性,同时对４种标记系统进行比较。在供试材料中筛选到具多态性的ＲＦＬＰ探针酶组合５６个,６６对ＳＳＲ引物,２０个ＲＡＰＤ引物和９个ＡＦＬＰ引物组合,分别检测到多态性带１６７、２０１、８７和１０８条。ＳＳＲ标记位点的平均多态性信息量（ＰＩＣ）最大（０．５４）,ＡＦＬＰ标记位点最小（０．３６）,但ＡＦＬＰ标记具有最高的多态性检测效率（Ａｉ,３２．２）。４种分子标记所得遗传相似系数相关性显著,比较相关系数表明 ＲＡＰＤ可靠性较低。依据 ４种分子标记结果将 １５个供试自交系划分为塘四平头、旅大红骨、兰卡斯特、瑞德和ＰＮ共５个类群,与系谱分析基本一致。认为ＳＳＲ和ＲＦＬＰ两种分子标记方法适合进行玉米种质遗传多样性的研究。     

A population genetics study of Anopheles darlingi (Diptera: Culicidae) from Colombia based on random amplified polymorphic DNA-polymerase chain reaction and amplified fragment lenght polymorphism markers

The genetic variation and population structure of three populations of Anopheles darlingi from Colombia were studied using random amplified polymorphic markers (RAPDs) and amplified fragment length polymorphism markers (AFLPs). Six RAPD primers produced 46 polymorphic fragments, while two AFLP primer combinations produced 197 polymorphic fragments from 71 DNA samples. Both of the evaluated genetic markers showed the presence of gene flow, suggesting that Colombian An. darlingi populations are in panmixia. Average genetic diversity, estimated from observed heterozygosity, was 0.374 (RAPD) and 0.309 (AFLP). RAPD and AFLP markers showed little evidence of geographic separation between eastern and western populations; however, the F ST values showed high gene flow between the two western populations (RAPD: F ST = 0.029; Nm: 8.5; AFLP: F ST = 0.051; Nm: 4.7). According to molecular variance analysis (AMOVA), the genetic distance between populations was significant (RAPD:phiST = 0.084; AFLP:phiST = 0.229, P < 0.001). The F ST distances and AMOVAs using AFLP loci support the differentiation of the Guyana biogeographic province population from those of the Chocó-Magdalena. In this last region, Chocó and Córdoba populations showed the highest genetic flow.     

AFLP analysis of the diversity and phylogeny of Lens and its comparison with RAPD analysis

AFLP and RAPD marker techniques have been used to evaluate and study the diversity and phylogeny of 54 lentil accessions representing six populations of cultivated lentil and its wild relatives. Four AFLP primer combinations revealed 23, 25, 52 and 48 AFLPs respectively, which were used to partition variation within and among Lens taxa. The results of AFLP analysis is compared to previous RAPD analysis of the same material. The two methods provide similar conclusions as far as the phylogeny of Lens is concerned. The AFLP technique detected a much higher level of polymorphyism than the RAPD analysis. The use of 148 AFLPs arising from four primer combinations was able to discriminate between genotypes which could not be distinguished using 88 RAPDs. The level of variation detected within the cultivated lentil with AFLP analysis indicates that it may be a more efficient marker technology than RAPD analysis for the construction of genetic linkage maps between carefully chosen cultivated lentil accessions.     

Genetic variation and population structure of oriental migratory locust, Locusta migratoria manilensis, in China by allozyme, SSRP-PCR, and AFLP markers

Allozyme analysis, microsatellite primer PCR (SSRP-PCR), and amplified fragment length polymorphism (AFLP) techniques were used to assess genetic diversity and population structure of the Chinese oriental migratory locust, Locusta migratoria manilensis. A total of 299 PCR markers (67 SSRPs and 232 AFLPs) were detected in eight populations, of which 98.7% were polymorphic markers. The proportion of polymorphic loci (95.5-98.8%) by SSRP+AFLP markers indicated no significant differences between populations, and all populations exhibited a similar level of variability; results of the allozyme analysis demonstrated that 19 loci gave rise to a lower level of polymorphism (55.6-66.7%). The genetic distances between the populations were relatively low. Shannon's index and Nei's gene diversity showed low differentiation among the populations. Allozyme analysis, however, reflected greater similarity and smaller differentiation between the populations than those shown by SSRP and AFLP markers. Neighbor-joining dendrograms derived from both the allozyme and SSRP+AFLP markers showed that the genetic distances among Chinese oriental migratory locust populations were not greatly influenced by geographic distance and breeding habitats.     

The comparison of RFLP,RAPD, AFLP and SSR (microsatellite) markers for germplasm analysis

The utility of RFLP (restriction fragment length polymorphism), RAPD (random-amplified polymorphic DNA), AFLP (amplified fragment length polymorphism) and SSR (simple sequence repeat, microsatellite) markers in soybean germplasm analysis was determined by evaluating information content (expected heterozygosity), number of loci simultaneously analyzed per experiment (multiplex ratio) and effectiveness in assessing relationships between accessions. SSR markers have the highest expected heterozygosity (0.60), while AFLP markers have the highest effective multiplex ratio (19). A single parameter, defined as the marker index, which is the product of expected heterozygosity and multiplex ratio, may be used to evaluate overall utility of a marker system. A comparison of genetic similarity matrices revealed that, if the comparison involved both cultivated (Glycine max) and wild soybean (Glycine soja) accessions, estimates based on RFLPs, AFLPs and SSRs are highly correlated, indicating congruence between these assays. However, correlations of RAPD marker data with those obtained using other marker systems were lower. This is because RAPDs produce higher estimates of interspecific similarities. If the comparisons involvedG. max only, then overall correlations between marker systems are significantly lower. WithinG. max, RAPD and AFLP similarity estimates are more closely correlated than those involving other marker systems.Abbreviations RFLP restriction fragment length plymorphism - RAPD random-amplified polymorphic DNA - AFLP amplified fragment length polymorphism - SSR simple sequence repeat - PCR polymerase chain reaction - TBE Tris-borate-EDTA buffer - MI marker index - SENA sum of effective numbers of alleles     

Characterisation of genetic diversity in potential biomass willows (Salix spp.) by RAPD and AFLP analyses.

The genus Salix (willow) contains a number of species which have great potential value as biomass crops in short rotation coppice (SRC). Efforts to improve biomass willows by breeding are currently hampered by the limited information available on genetic diversity and on genetic relationships within and among species, clones, and hybrids in the gene pool. Hybridisation occurs commonly in nature and the relatedness of many clones is unclear. Molecular markers were used to assess genetic diversity in a reference set of willows maintained within the U.K. National Collection and 16 elite clones currently being evaluated in field trials at several European sites. The two marker systems tested, RAPDs and AFLPs, were equally informative for revealing relationships within the reference set of clones. No differences were observed when alternative similarity coefficients were compared or when analysis was restricted to the use of polymorphic bands only. Good agreement with available knowledge of the clonal origins was obtained and one instance of duplicate clones was identified. AFLPs revealed more genetic diversity and discriminated between closely related clones. A difference in the relationships revealed was observed with one AFLP primer combination. RAPDs were more problematic, both in terms of reproducibility and scorability.     

Comparison of PCR-based marker systems for the analysis of genetic relationships in cultivated potato

The application of AFLPs, RAPDs and SSRs to examine genetic relationships in the primary northwestern European cultivated potato gene pool was investigated. Sixteen potato cultivars were genotyped using five AFLP primer combinations, 14 RAPD primers, and 17 database-derived SSR primer pairs. All three approaches successfully discriminated between the 16 cultivars using a minimum of one assay. Similarity matrices produced for each marker type on the basis of Nei and Li coefficients showed low correlations when compared with different statistical tests. Dendrograms were produced from these data for each marker system. The usefulness of each system was examined in terms of number of loci revealed (effective multiplex ratio, or EMR) and the amount of polymorphism detected (diversity index, or DI). AFLPs had the highest EMR, and SSRs the highest DI. A single parameter, marker index (MI), which is the product of DI and EMR, was used to evaluate the overall utility of each marker system. The use of these PCR-based marker systems in potato improvement and statutory applications is discussed.Abbreviations: PCR, polymerase chain reaction; AFLP, amplified fragment length polymorphism; RAPD, randomly amplified polymorphic DNA; DNA, deoxyribonucleic acid; EMR, effective multiplex ratio; DI, diversity index; MI, marker index; RFLP, restriction fragment length polymorphism.