SDS-PAGE蛋白电泳技术在F1代杂交种纯度鉴定中的应用

随着市场的需要,商品种子量越来越大,而对杂交种质量问题不容忽视,我们在实际工作中对中椒四号,中椒五号,京欣一号,金花宝四个杂交种纯度鉴定尝试了大量的同工酶电泳方法,包括ＰＯＸ,ＥＳＴ,ＡＤＨ,ＭＤＨ,ＩＳＨ,ＰＧＭ,ＰＧＩ等同工酶电泳图谱中均未找到交父母本与一代杂种的鉴别标记。而在种子蛋白质ＳＤＳ－ＰＡＧＥ电泳图谱中找到了一代杂种与父母本的鉴别带,并利用这个鉴别带进行了杂交种纯度鉴定。这种鉴定方法的准确性在田间对照纯度鉴定中得到确认,此方法可用于杂交种商品种子的纯度检测。     

利用RAPD技术鉴定大白菜主栽品种及品种间遗传多样性分析

采用随机引物扩增多态ＤＮＡ（ＲＡＰＤ）技术分析了２１个大白菜主栽品种。用１３个引物共扩增出８７个清晰可重复的ＤＮＡ片段,其中３９条带具有多态性,多态性条带的频率为４４．８％。ＯＰＥ０１是多态性频率最高的引物,它可区分１５个大白菜品种,再与引物ＯＰＨ０３和ＯＰＨ１２配合,即可将２１个大白菜品种区别开。     

用RAPD方法鉴定杂交辣椒种子纯度

本文用ＲＡＰＤ（随机扩增多态性ＤＮＡ）方法对辣椒种子的父本、母本及杂种一代进行分析研究,筛选出适合辣椒杂种一代种子纯度鉴定的随机引物Ｓ５,并对并进了种子纯度的初步鉴定。ＲＡＰＤ结果和田间鉴定的相比,在误差允许范围内,进一步证明了ＲＡＰＤ方法在蔬菜种子纯度鉴定中应用的可行性。     

‘福春1号’大白菜一代杂种纯度的SRAP鉴定

‘福春1号’大白菜是福州市蔬菜科学研究所选育的晚抽薹春白菜新品种。为鉴定该品种的纯度,利用80对SRAP引物组合对‘福春1号’及其亲本混合池DNA模板进行扩增,筛选出杂交种具有亲本互补特征带的引物组合,并利用该组合对田间栽培的‘福春1号’大白菜幼苗进行纯度检测。本研究最终筛选出引物组合M2+E4,其对杂交种扩增产生亲本互补的特征带,可将亲本和杂交种有效区分。利用这对SRAP引物组合对102株‘福春1号’大白菜幼苗进行纯度检测,检测纯度为94.12%。SRAP标记鉴定结果与田间调查结果基本一致。SRAP分子标记技术可用于‘福春1号’大白菜的纯度鉴定。     

黄瓜‘浙秀1号’种子纯度的SSR鉴定

种子纯度在种子生产中的重要性日益提升,分子标记技术因其具有高效、稳定、可靠等优点越来越多地被应用到种子纯度鉴定中。本研究利用SSR分子标记技术对黄瓜杂交品系‘浙秀1号’及其两亲本之间的多态性进行了引物筛选和种子纯度鉴定研究。结果表明,在699对供选SSR引物中,有3对引物分别在‘浙秀1号’杂交种和其双亲之间表现为稳定的共显性,杂交种带型为双亲的互补带型,适合于杂交种纯度鉴定。经田间试验验证,引物稳定性良好,且与田间鉴定结果非常接近,可用于‘浙秀1号’杂交种种子纯度的鉴定,显示出SSR标记技术在黄瓜杂交种子纯度的室内快速检测中有广泛应用前景。     

小麦株高近等基因系的RAPD标记研究

采用ＲＡＰＤ技术，以４个小麦株市基因近等基因系（分别含有rht、Ｒht1、Rht2、Rht3)为材料，对２９６个单一随机引物（１０个核苷酸）进行了筛选。发现２５个引我的扩增产物的近等基因系间表现出特异性，在６次重复试验中，有１８个引物的特异护增片段不能重复，６个引物可以重复２－３资，唯有ＯＰＡＭ０１在全部试验中均能稳定重复，其特异扩增版段ＯＰＡＭ０１１８６０可以作为rht基因的ＲＡＰＤ标记。     

利用SSR标记鉴定茎瘤芥(榨菜)“涪杂5号”种子纯度

为建立一套丰产抗病型的茎瘤芥杂交品种"涪杂5号"种子纯度的快速鉴定方法,本研究以"涪杂5号"及其双亲为试验材料,采用改良CTAB法快速提取DNA与SSR标记分析相结合,对茎瘤芥杂交种种子纯度进行检测。利用改良CTAB法提取的DNA能够满足SSR分析要求,从600对SSR芸薹属共有引物中筛选出3对共显性SSR标记引物Ol11-G11、Na14-G06和Na14-G10,父母本条带互补,可将杂交种中的假杂株分离开。分别利用其中两对引物相结合,对"涪杂5号"种子群体进行SSR检测,结果发现分子标记鉴定结果与田间鉴定结果基本一致。研究结果表明了利用SSR引物Na14-G06与Na14-G10或Ol11-G11相结合可实现对茎瘤芥杂交种"涪杂5号"种子纯度的快速和准确鉴定,并为其大面积推广和安全生产提供了理论依据。     

SSR标记对宁单11种子纯度的鉴定

采用SSR分子标记方法,选用24对引物,以杂交玉米宁单11杂交种子及其父母本为实验材料,对宁单11种子纯度进行鉴定。通过带型分析,筛选出在杂交种中呈现清晰且双亲互补带型的特异性引物,并依据特异性引物的筛选标记结果对抽样批次的宁单11种子样本进行纯度鉴定。结果表明:phi034、phi080、umc1638、bnlg439、bnlg1792五对SSR引物可用于玉米杂交种宁单11种子纯度鉴定。     

分子标记方法鉴定玉米品种纯度的应用试验

利用SSR分子标记技术,选用15对SSR引物以玉源5号玉米杂交种及其亲本为材料,筛选出了3对在杂交种中呈现双亲互补带型的引物,并在玉源5号杂交种中人为混入双亲,通过本研究确定的方法和引物,可准确鉴定出混入的自交系。研究表明:这3对引物均可用于玉米杂交种玉源5号种子的纯度鉴定。     

利用SSR标记鉴定玉米杂交种黔单16号纯度的研究

玉米杂交种纯度鉴定对玉米种子质量管理具有重要意义;本实验利用SSR标记技术,以玉米杂交种黔单16号及其亲本DNA为供试材料,从30对SSR引物中筛选出3对在杂交种黔单16号中呈现双亲带型的引物;研究表明,这3对引物均可用于玉米杂交种黔单16号种子纯度鉴定.     

Purity control of F1-hybrid tomato cultivars by RAPD markers

Randomly amplified polymorphic DNA (RAPD) markers were applied in purity control of hybrid seed production of tomato (Lycopersicon esculentum Mill.). DNA from three commercial F₁-hybrid cultivars and their parental lines was subjected to RAPD screening with 50 primers. Two of four primers which detected polymorphism between the parents tested, generated paternal-specific RAPDs, enabling a clear distinction to be made between hybrids and their maternal parents. In addition, combination of the polymorphic DNA products generated by these primers exhibited hybrid-specific patterns, enabling each cultivar to be identified. This result indicates the practical usefulness of RAPD markers in hybrid-tomato-seed purity-control tests and cultivar identification. The approach is advantageous in its rapidity and simplicity, particularly as an alternative for those cultivars for which lengthy and costly phenotypic tests are currently used.     

甘蓝型油菜中油杂8号种子纯度的SSR鉴定

（中国农业科学院油料作物研究所/国家油料作物改良中心/农业部油料作物遗传改良重点实验室,湖北武汉 430062）     

杂交油葵A15种子纯度的RAPD鉴定

从杂交油葵A15及其亲本中提取基因组DNA,用180个RAPD随机引物进行扩增,从中筛选出3个可将亲本和子代区分开的引物OPD09、OPD12和OPK12。OPD09产生亲本互补的特征带OPD09-1470bp、OPD09-870bp;OPD12产生母本特征带OPD12-1230bp,OPK12产生父本特征带OPK12-1540bp、OPK12-940bp,上述谱带均在子代中出现。以单引物(OPD09)和双     

Varietal identification in Cichorium intybus L. and determination of genetic purity of F1 hybrid seed samples, based on RAPD markers

Random amplified polymorphic DNA (RAPD) markers were used to distinguish between several Cichorium intybus genotypes, comprising four white witloof inbred lines, three red witloof experimental inbred lines and a number of F1 hybrids derived from two white parents. Amplification conditions and reproducibility of RAPD patterns were examined. Comparison of polymerase chain reaction (PCR) products obtained by using 100 10-mer arbitrary primers allowed identification of all the lines analysed. With several primers, we defined line-specific RAPD markers, while with others polymorphism was more extensive, revealing several RAPD markers for several lines. All the differences were confirmed both on individual heads and young seedlings for each genotype. Because of the Mendelian segregation of these molecular markers, this method was applied to evaluate the genetic purity of F1 hybrid seed samples.     

Assessment of DNA markers for seed contamination testing and selection of disease resistance in cabbage

Cabbage (Brassica oleracea L. var. capitata) is an important vegetable worldwide. Most Japanese commercial cultivars of cabbage use an F₁ hybrid seed production system. The purity of F₁ hybrid seeds is important and the assessment of purity based on DNA markers can be highly accurate. In addition, selection of agronomically important traits such as disease resistance based on DNA markers is useful for breeding of cabbage. The aim of this study is to demonstrate the effectiveness of DNA marker-assisted selection in cabbage. In this study we distinguished the parental S haplotypes in 35 F₁ hybrid cultivars by combining several linked DNA markers. Thirty-one highly polymorphic simple sequence repeats (SSR) markers were screened from 175 reported SSR markers, which are useful for assessment of the purity of F₁ hybrid seeds. We examined the relationship between the DNA marker based genotype and the phenotype by an inoculation test of clubroot disease. A co-dominant PCR–RFLP marker was developed for selection of Fusarium yellows resistance and the genotypes using this marker were consistent with inoculation test in all tested samples.     

EST-SSR marker-based assay for the genetic purity assessment of safflower hybrids

Full potential of any hybrid can be exploited by ensuring the supply of genetically pure seeds. Conventionally, hybrid seed purity assessment is done through grow out test (GOT) which is based on the morphological and floral characters of plants grown to maturity. Being land and labor intensive, time consuming and influenced by the environment, there is an immense need to replace GOT with a simple, rapid, unbiased and cost-effective DNA based assay for hybrid purity assessment. With this objective, the parental lines of three commercial safflower hybrids of India (NH-1, NH-15 and DSH-129) were screened using 74 safflower EST-SSR markers and five markers were found to be polymorphic. A PCR-based assay with these markers showed both alleles of the parental lines in pure hybrids proving the heterozygosity, while the off-types were identified by the presence of either of the parental alleles. This assay could accurately determine the genetic purity in a predetermined sample of hybrids constituted by deliberately mixing seeds of parental lines. This is the first report demonstrating the utility of EST-SSR markers for the assessment of genetic purity of hybrids in crop plants.     

同工酶差异位点分析在蔬菜杂交种纯度检测中的应用

用10种同工酶和蛋白质分析体系，分析了7种重要蔬菜的71个杂交种与其亲本之间的差异位点，以及这些差异位点用于杂交种纯度检测的可能性和存在的问题。试验表明，作物的不同种类，杂交种与亲本之间的亲缘关系以及作物各类的遗传多态性，都会影响同工酶差异位点产生的多寡，从而影响到这一技术是否能用于此种作物的纯度检测。分析了不同同工酶在不同蔬菜中的多态性以及在蔬菜种子纯度检测中的表现。     

RAPD技术快速鉴定辣椒杂种纯度的研究

利用RAPD技术，通过对60个随机引物的筛选，获得能区分粤椒4号及其亲本的引物OPS—01、并得到单株验证。     

白菜类蔬菜种子纯度SSR分子标记鉴定

种子纯度是种子质量的核心指标,是保障优良品种增产的关键因素。为了鉴定白菜种子纯度,本研究利用72对SSR分子标记检测种子纯度并结合田间植株形态观察,鉴定了5个白菜类优势品种的种子纯度,分别是‘珍绿6号’‘、津夏3号’‘、津研快绿1号’‘、速俊228’‘、速俊316’。结果表明,从72对SSR引物中筛选出5对具有多态性的引物,能够将父母本与杂交种区分开。试验中SSR分子鉴定结果虽略低于田间形态学鉴定结果,但利用统计学进行显著性分析,发现两者间无显著性差异。说明SSR分子标记可以用于白菜种子纯度的鉴定并通过本研究建立了一套准确、有效的品种纯度鉴定体系。