巴马小型猪群体遗传结构的随机扩增多态DNA分析

目的分析巴马小型猪的群体遗传结构。方法应用经过筛选的 ３１条引物对巴马小型猪个体基因组ＤＮＡ进行ＲＡＰＤ扩增,利用从ｉｎｔｅｒｎｅｔ网上下载的软件ＲＡＰＤ ｉｓｔａｎｃｅ Ｐａｃｋａｇｅ ｖｅｒ－ｓｉｏｎ１．０４软件对实验结果进行分析,计算不同个体间的相似系数。结果经ＲＡＰＤ反应,共扩增出２７５条带,其中多态性带８５条,多态性带频率在０～６６．７％之间,平均为２７．７％。不同个体猪拥有的ＲＡＰＤ条带具有差异,但拥有相同条带个体猪比率较高。该群体猪的相似系数（Ｆ）为０．９２８（０．７８～０．９７）,平均等位基因频率（ｑ）为０．７３２,最低平均杂合率（Ｈ）为０．２８６。结论巴马小型猪个体具有较好的遗传一致性和遗传稳定性。     

辣椒疫霉产毒共分离RAPD标记的研究

辣椒疫毒（Ｐｈｙｔｏｐｈｔｈｏｒａ ｃａｐｓｉｃｉ）毒素的产生是由一个不完全显性基因所控制,通过ＲＡＰＤ／ＢＳＡ分析证明,用ＯＰＷ１２扩增得到一条约１３００ｂｐ的ＲＡＰＤ标记带,该带与辣椒疫霉产毒共分离。纯化回收ＯＰＷ１２１３００ＤＮＡ,共转化感受态Ｅ．ｃｏｌｉ ＤＨ５α,筛选出３个白色阳性克隆,序列分析发现该标记ＤＮＡ为１２９１ｂｐ。这一标记ＤＮＡ序列的阐明为进一步分析产毒遗传机理提供了新的信息     

用RAPD和AFLP的方法对中国卤虫（Autemia）种及亲缘关系的研究

利用ＲＡＰＤ（随机扩增多态ＤＮＡ）和ＡＦＬＰ（扩增片段长度多态性）技术对不同种及种群卤虫的关系进行分析。１０１个随机引物对４种卤虫Ａ．ｆｒａｎｃｉｓｃａｎａ、Ａ．ｕｒｍｉａｎａ、Ａ．ｓｉｎｉｃａ和Ａ．ｐａｒｔｈｅｎｏｇｅｎｅｔｉｃａ基因组ＤＮＡ进行扩增,平均每个种获得７５１条带,其中４５８条带为多态性标记,每个引物提供平均７．４个标记信息,聚类结果表明Ａ．ｓｉｎｉｃａ是不同于其他旧大陆两性生殖卤虫     

RAPD标记构建水稻分子连锁图

利用随机扩增多态性ＤＮＡ（ＲＡＰＤ）,在一个水稻（Ｏｒｙｚａ ｓａｔｉｖａ Ｌ．）的双单倍体（ＤＨ）群体中发展分子标记,仅用５２ 个ＲＡＰＤ标记建成了一个水稻ＲＡＰＤ分子连锁图。该图覆盖基因组的总长度为８９８．４ ｃＭ （ｃｅｎｔｉｍ ｏｒｇａｎ）,标记间的平均间距为１７．３ ｃＭ,它能与用同一群体构成的ＲＦＬＰ图谱互相补充     

辣椒疫霉产毒共分离RAPD标记的研究

辣椒疫霉（Ｐｈｙｔｏｐｈｔｈｏｒａ　ｃｏｐｓｉｃｉ）毒素的产生是由一个不完全显性基因所控制,通过ＲＡＰＤ／ＢＳＡ分析证明,用ＯＰＷ１２扩增得到一条约１３００ｂｐ的ＲＡＰＤ标记带,该带与辣椒疫霉产毒共分离。纯化回收ＯＰＷ１２　１３００ＤＮＡ,共转化感受态Ｅ．ｃｏｌｉ　ＤＨ５ａ,筛选出３个白色阳性克隆,序列分析发现该标记ＤＮＡ为１２９１ｂｐ。这一标记ＤＮＡ序列的阐明为进一分析产毒遗传机理提供了新的信息。     

辣椒疫霉产毒共分离RAPD标记的研究*

辣椒疫霉（Ｐｈｙｔｏｐｈｔｈｏｒａ　ｃｏｐｓｉｃｉ）毒素的产生是由一个不完全显性基因所控制,通过ＲＡＰＤ／ＢＳＡ分析证明,用ＯＰＷ１２扩增得到一条约１３００ｂｐ的ＲＡＰＤ标记带,该带与辣椒疫霉产毒共分离。纯化回收ＯＰＷ１２　１３００ＤＮＡ,共转化感受态Ｅ．ｃｏｌｉ　ＤＨ５ａ,筛选出３个白色阳性克隆,序列分析发现该标记ＤＮＡ为１２９１ｂｐ。这一标记ＤＮＡ序列的阐明为进一分析产毒遗传机理提供了新的信息。     

采用PCR—RFLP和RAPD对球壳孢目真菌系统学的研究

对球壳孢目真菌首次采用ＰＣＲ－ＲＦＬＰ和ＲＡＰＤ进行了系统发育研究,以ＩＴＳ１和ＩＴＳ４为引物对４属１２种２４个菌株的核糖体ＤＮＡ转录间区进行了ＰＣＲ扩增,４种内切酶酶切,结果表明：主要属的ＩＴＳ区长度不同,同属不同种相同。Ｃｏｎｉｏｔｈｙｒｉｕｎ,Ｐｈｙｌｌｏｓｔｉｃｔａ,Ａｓｃｏｃｈｙｔａ,ＳｅｔｏｒｉａＩＴＳ区长度分别为６３０,５６０,５５０,５４０ｂｐ;酶切图各间差别明显,属内种间基本一致     

黑麦1R染色体的微切微克隆研究

显微分离出黑麦（ＳｅｃａｌｅｃｅｒｅａｌｅＬ．）１Ｒ染色体,用ＣｏｈｅｓｉｖｅａｄａｐｔｅｒｓｓｉｎｇｌｅｐｒｉｍｅｒＰＣＲ（ＣＡＳＰＰＣＲ）方法进行体外扩增,以ＤＩＧ１１ｄＵＴＰ标记扩增产物为探针,进行Ｓｏｕｔｈｅｒｎ分子杂交,结果表明扩增产物来自黑麦１Ｒ染色体。用１／１０体积的连接物转化Ｅ．ｃｏｌｉＤＨ５α,获得１００００多个重组菌落。经酶切分析,克隆子的插入片段为２５０～５００ｂｐ,为进一步筛选１Ｒ染色体的分子标记打下了基础     

RAPD技术及其在微生物学方面的应用

198 0年 ,Ｂｏｔｓｅｉｎ提出ＤＮＡ限制性片段长度多态性 (ＲＦＬＰ)可以作为遗传标记 ,从此开创了直接应用ＤＮＡ多态的新阶段。 80年代后 ,ＤＮＡ多聚酶链式反应 (ＰＣＲ)的发展 ,使直接扩增ＤＮＡ的多态性成为可能 ,并在此基础上产生了许多种新型分子标记 ,诸如扩增片段多态性 (ＡＬＦＲ)、串联重复序列(ＶＮＴＲ)、单链构型多态性 (ＰＣＲ ＳＳＣＰ)、序列特异扩增区域 (ＳＣＡＲ)、随机扩增多态性ＤＮＡ(ＲＡＰＤ)等。而ＲＡＰＤ是较为突出的一种。ＲＡＰＤ是由Ｗｉｌｌｉａｍｓ和Ｗｅｌｓｈ在 1 990年各自独立发现的一种ＤＮＡ多态检…     

利用分子标记定位农垦58S的光敏核不育基因

对农垦５８Ｓ（Ｏｒｙｚａｓａｔｉｖａｓｐ．ｊａｐｏｎｉｃａ）／大黑矮生标记基因系ＦＬ２组合组建可育集团和不育集团,并以亲本为对照进行了ＲＦＬＰ、ＲＡＰＤ和双引物ＲＡＰＤ分析,结果第１２染色体上的一个单拷贝标记Ｇ２１４０与光敏核不育基因连锁遗传,二者间的遗传图距为１４．１ｃＭ（ｃｅｎｔｉｍｏｒｇａｎ）。在筛选过的１０４０个随机单引物和１９０个双引物中,仅引物ＯＰＡＵ１０扩增出与光敏核不育基因连锁的１．５ｋｂＤＮＡ片段,回收、克隆该ＤＮＡ片段并制备探针,将其转换成共显性的ＲＦＬＰ标记并命名为ＯＰＡＵ１０１５００。分离群体连锁分析表明该标记与标记Ｇ２１４０紧密连锁,将农垦５８Ｓ的一对光敏核不育基因定位于第１２染色体上。     

Molecular genetic marker-based approaches to the verification of lilac Syringa vulgaris L. in vitro germplasm collections

RAPD analysis was used to verify the varieties in an in vitro germplasm collection of lilac Syringa vulgaris L. RAPD patterns were obtained with 16 decanucleotide primers for 46 accessions (microclones and corresponding reference varieties). The RAPD patterns of a microclone and the corresponding reference variety often differed in composition; consequently, it was infeasible to verify the accessions by direct comparison of the RAPD patterns. Hence, evaluation of the relative genetic distances between accessions (microclones) and known varieties was proposed as a method to verify lilac in vitro germplasm collections.     

Polymorphic glutamate dehydrogenase in lilac vitroplants as revealed by combined preparative IEF and native PAGE: Effect of ammonium deprivation, darkness and atmospheric CO2 enrichment upon isomerization

The activity and polymorphism of glutamate dehydrogenase (GDH) were studied in basal callus of lilac ( Syringa vulgaris L.) vitroplants. Native PAGE alone revealed seven bands staggered at regular intervals. Preparative liquid-vein IEF allowed the separation of six to ten GDH fractions with charges ranging between 5.18 and 7.08. Analysis of these GDH fractions in native PAGE indicated that up to seven GDH bands can be detected for each fraction. This suggests the existence of seven isoforms of the enzyme with subunits presenting different isoelectric points. Dark- and ammonium-controlled forms were found to be the more acidic and faster migrating ones in native PAGE. The results support for the first time that atmospheric CO₂ enrichment increases GDH activity dramatically and modifies isomerization of the enzyme.     

RAPD分析花椰花不同品种的遗传变异

利用随机扩增多态性DNA（RAPD）方法,对13个花椰花品种的基因组DNA进行多态性分析。选用20个10bp随机引物,共扩增出175条DNA片段,其中多态性片段118例,占67．4％,结果表明,花椰花品种间具有丰富的遗传多样性。依据扩增结果进行遗传相似系数分析,构建聚类分析树状图。初步探讨了各品种间的遗传变异关系及RAPD技术在花椰花种质资源分类鉴定和育种工作中的应用前景。     

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0.26kb-1.98kb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100bp-500bp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0．6495,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0．6884。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。     

Qualitative and quantitative characterization of RAPD variation among snap bean (Phaseolus vulgaris) genotypes

Ten snap bean (Phaseolus vulgaris) genotypes were screened for polymorphism with 400 RAPD (random amplified polymorphic DNA) primers. Polymorphic RAPDs were scored and classified into three categories based on ethidium bromide staining intensity. An average of 5.19 RAPD bands were scored per primer for the 364 primers that gave scorable amplification products. An average of 2.15 polymorphic RAPDs were detected per primer. The results show that primer screening may reduce the number of RAPD reactions required for the analysis of genetic relationships among snap-bean genotypes by over 60%. Based on the analysis of the distribution of RAPD amplification, the same number of polymorphic RAPDs were amplified from different genotypes for all RAPD band intensity levels. A comparison of RAPD band amplification frequency among genotypes for the three categories of bands classified by amplification strength revealed a measurable difference in the frequencies of RAPDs classified as faint (weakly amplifying) compared to RAPD bands classified as bold (strongly amplifying) indicating a possible scoring error due to the underscoring of faint bands. Correlation analysis showed that RAPD bands amplified by the same primer are not more closely correlated then RAPD bands amplified by different primers but are more highly correlated then expected by chance. Pairwise comparisons of RAPD bands indicate that the distribution of RAPD amplification among genotypes will be a useful criterion for establishing RAPD band identity. For the average pairwise comparison of genotypes, 50% of primers tested and 15.8% of all scored RAPDs detected polymorphism. Based on RAPD data Nei's average gene diversity at a locus was 0.158 based on all scorable RAPD bands and 0.388 if only polymorphic RAPD loci were considered. RAPD-derived 1 relationships among genotypes are reported for the ten genotypes included in this study. The data presented here demonstrate that many informative, polymorphic RAPDs can be found among snap bean cultivars. These RAPDs may be useful for the unique identification of bean varieties, the organization of bean germplasm, and applications of molecular markers to bean breeding.     

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.     

遗传改良"新吉富" (NEW GIFT)尼罗罗非鱼RAPD-SCAR标记开发及其在品系鉴别中的应用

“新吉富”(NEW GIFT)尼罗罗非鱼是以1994年引进的 “吉富”(GIFT)品系尼罗罗非鱼为基础群体,经过14年9代系统选育后获得的新品种,该品种已被国家审定为良种,已在全国推广,迫切需要有效的分子遗传标记来鉴别该新品种并实施科学管理。该研究对良种群体开展了RAPD分析,并进一步将特异性的RAPD标记转化成了稳定的SCAR标记。 通过对“新吉富”群体的RAPD分析,找到了两个品种特异性的RAPD条带（S₃₀₄^{624 bp}和S₃₆^{568 bp}）。对品种特异性的RAPD条带进行了胶回收、克隆和序列测定。根据测序结果设计了两对位点特异性引物来扩增这两个品种特异性条带（简称SCAR标记Ⅰ和Ⅱ）,检测这两个SCAR标记在“新吉富”、“吉富”以及国内外7个养殖品系中的出现频率。SCAR标记Ⅰ（553 bp）在“新吉富”群体中的出现频率为85.7%,而在其基础群体（“吉富”）中的出现频率仅为16.7%;SCAR标记Ⅱ（558 bp）在“新吉富”群体中的出现频率达到91.4%,但在其它7个养殖品系中的出现频率仅在0%～70%之间。为验证这两个标记的可靠性,检测了这两个标记在一个埃及罗非鱼野生群体中的出现频率,发现SCAR标记Ⅰ和SCAR标记Ⅱ的出现频率分别为10%和70%,远低于在“新吉富”群体中的出现频率。这两个SCAR标记在“新吉富”良种群体中的高出现频率,预示它们可能与生长性能相关QTL位点间存在连锁关系,显示SCAR标记技术对选育世代间的纵向追溯、以及选育良种群体与其它养殖品系间的横向鉴别有较好应用前景。     

用RAPD分析多年生黑麦草品种间遗传多样性

多年生黑麦草是世界上应用最广泛的冷季型草坪草兼牧草之一,具有建植快速,分蘖能力强,适口性良好等特点。为了评价其品种间的遗传多样性,选取了多年生黑麦草的14个草坪型品种和2个意大利黑麦草品种用RAPD进行遗传多样性分析,并且对每个品种的几项坪用特性进行了观察。61个引物共扩增出408条带,多态条带比率(PPB)为89．95％。聚类分析结果表明,多年生黑麦草品种之间的遗传多样性较低,相似系数分布于44．55％～95．69％之间。一年生黑麦草的2个品种TopOne和A,vance没有单独聚成一类,而是与14个多年生黑麦草品种混合聚在一起,而且Avance与其他多年生黑麦草之间的相似系数较高。聚类结果与形态特征没有明确的对应关系。     

Evaluation of genetic diversity and uniformity of head cabbage DH lines by the use of RAPD markers

DH lines derived from cabbage cvs. Kamienna G?owa, S?awa z Enkhuizen and Langendijker, representing R1 generation, were analysed by the use of RAPD markers for their diversity and uniformity. For the evaluation of genetic diversity, eight primers yielding informative bands were used. Of the total of 83 RAPD bands scored in this study, 16.9% were polymorphic between a set of 13 DH lines. The similarity of the DH lines, estimated by Jaccard's coefficient, was depicted in the UPGMA dendrogram. Fourteen generated informative RAPD bands allowed the identification of DH lines developed from each cultivar. The evaluation of the uniformity for six closely related DH lines was possible by the use of three primers which generate one or two polymorphic bands. The lack of differences among ten plants of the five investigated DH lines manifested their uniformity. One line showed intraline polymorphism with two RAPD primers. The occurrence of the differences at the molecular level among ten plants indicated that their parental R0 plant was probably obtained from somatic cells, not by androgenesis.     

A comparative analysis of genetic diversity in blackgram genotypes using RAPD and ISSR markers

Random amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR) markers were used to study the DNA polymorphism in elite blackgram genotypes. A total of 25 random and 16 ISSR primers were used. Amplification of genomic DNA of the 18 genotypes, using RAPD analysis, yielded 104 fragments that could be scored, of which 44 were polymorphic, with an average of 1.8 polymorphic fragments per primer. Number of amplified fragments with random primers ranged from two (OPA-13) to nine (OPK-4) and varied in size from 200 bp to 2,500 bp. Percentage polymorphism ranged from 16.6% (OPK-7) to a maximum of 66.6% (OPE-5, OPH-2, and OPK-8), with an average of 42.7%. The 16 ISSR primers used in the study produced 101 bands across 18 genotypes, of which 55 were polymorphic. The number of amplified bands varied from two (ISSR 858) to ten (ISSR 810), with a size range of 200–2,200 bp. The average numbers of bands per primer and polymorphic bands per primer were 6.3 and 3.4, respectively. Percentage polymorphism ranged from 25% (ISSR 885) to 100% (ISSR 858), with an average percentage polymorphism of 57.5% across all the genotypes. The 3-anchored primers based on poly(GA) and poly(AG) motifs produced high average polymorphisms of 54.98% and 58.32%, respectively. ISSR markers were more efficient than the RAPD assay, as they detected 57.4% polymorphic DNA markers in Vigna mungo as compared to 42.7% for RAPD markers. The Mantel test between the two Jaccards similarity matrices gave r =0.32, showing low correlation between RAPD- and ISSR-based similarities. Clustering of genotypes within groups was not similar when RAPD and ISSR derived dendrogram were compared, whereas the pattern of clustering of the genotypes remained more or less the same in ISSR and combined data of RAPD and ISSR.