RAPD analysis of genetic variation in natural populations of Betula alnoides from Guangxi, China

There is an urgent need for the developmentof early identification techniques inolive-trees due to the economic importanceof cultivar identification in periods ofexpansion like now. We have been able toidentify 22 olive-tree cultivars using only10 different, specific, repeatable markers.These markers were designed by the cloningof significant RAPD bands obtained in PCRperformed on bulked DNA to retain thegenetic variability of each cultivar.Clones were partially or totally sequencedand new primers derived from thesesequences were used to obtain SequenceCharacterised Amplified Region (SCAR)fragments. We have demonstrated that theuse of the 10 SCAR markers is enough toprovide a simple, cheap, and reliableprocedure to identify 22 geographicallyrelated olive-tree cultivars.     

Novel male-specific molecular markers (MADC5, MADC6) in hemp

Decamer RAPD primers were tested on dioecious and monoecious hemp cultivars to identify sex-specific molecular markers. Two primers (OPD05 and UBC354) generated specific bands in male plants. These two DNA fragments were isolated, cloned and sequenced. Both markers proved to be unique, since no sequence with significant homology to OPD05₉₆₁ and UBC354₁₅₁ markers were found in databases. These markers were named MADC3 (OPD05₉₆₁) and MADC4 (UBC354₁₅₁) (Male-Associated DNA from Cannabis sativa). The markers were converted into sequence-characterized amplified region (SCAR) markers. The SCAR markers correlated with the sex of the segregating F₂ population and proved the tight linkage to the male phenotype. Results of F₂ plant population analysis suggest these markers are to be linked to the Y chromosome. This revised version was published online in August 2006 with corrections to the Cover Date.     

Identification of a molecular marker linked to an Agropyron elongatum-derived gene Lr19 for leaf rust resistance in wheat

The leaf rust resistance gene Lr19, transferred from Agropyron elongatum into wheat (Triticum aestivum L.) imparts resistance to all pathotypes of leaf rust (Puccinia recondita f.sp. tritici) in South‐east Asia. A segregating F₂ population from a cross between the leaf rust resistant parent ‘HW 2046’ carrying Lr19 and a susceptible parent ‘Agra Local’ was screened in the phytotron against a virulent pathotype 77‐5 of leaf rust with the objective of identifying the molecular markers linked to Lr19. The gene was first tagged with a randomly amplified polymorphic DNA (RAPD) marker S73₇₂₈. The RAPD marker linked to the gene Lr19 which mapped at 6.4 ± 0.035 cM distance, was converted to a sequence characterized amplified region (SCAR) marker. The SCAR marker (SCS73₇₁₉) was specific to Lr19 and was not amplified in the near‐isogenic lines (NILs) carrying other equally effective alien genes Lr9, Lr28 and Lr32 enabling breeders to pyramid Lr19 with these genes.     

Resistance to Tomato spotted wilt virus introgressed from Lycopersicon peruvianum in line UPV 1 may be allelic to Sw-5 and can be used to enhance the resistance of hybrids cultivars

The breeding line UPV 1 developed from the PE-18 accession of Lycopersicon peruvianum collected in Huallanca, Ancash, Peru, shows resistance to TSWV. Mechanical inoculation and thrips transmission were used to study the inheritance of TSWV resistance of this line. UPV 1resistance is controlled by a dominant gene. The penetrance of this resistance gene was complete in mechanical inoculation and incomplete when thrips transmission was used. Linkage tests between the resistance genes of lines UPV 1 and RDD (Sw-5), indicated allelism. A molecular analysis using a SCAR marker tightly linked to Sw-5 also supported this hypothesis. In heterozygotes the level of resistance expressed in UPV 1 is higher than that expressed in RDD (Sw-5), indicating that the resistance from UPV 1 may be of higher value for the development of commercial hybrids. This revised version was published online in July 2006 with corrections to the Cover Date.     

SSR and SCAR markers linked to the fertility-restoring gene for a D2-type cytoplasmic male-sterile line in wheat

‘Yi 4060’ is an elite restorer line of a non‐photoperiod‐sensitive D²‐type cytoplasmic male‐sterile (CMS) line of wheat. Random amplified polymorphic DNA (RAPD) and simple sequence repeat (SSR) markers were employed to map one major fertility‐restoring gene (D²Rf₁) in ‘Yi 4060′. The sterile and fertile DNA pools were established from individuals in BC₆, based on bulked segregant analysis. One RAPD marker E09, linked to D²Rf₁, was converted to a SCAR marker and designated as E09‐SCAR₈₆₅. The genetic distance between E09‐SCAR₈₆₅ and D²Rf₁ is 9.5 cM. Two SSR markers, Xgwm11 and Xgwm18, were also linked to a D²Rf₁ and co‐segregated with E09‐SCAR₈₆₅. The three molecular markers are useful in marker‐assisted breeding of the elite restorer lines for D² ‐type CMS lines in wheat.     

小麦秆锈菌生理小种21C3CTH SCAR标记的建立

为了建立中国小麦秆锈菌流行小种的分子检测标记,利用RAPD(Random amplified polymorphic DNA)技术对我国小麦秆锈菌6个主要生理小种21C3CTH、21C3CPH、21C3CFH、34MKG、34C2MKK和34C2MKR进行了分析,筛选特异性片段并将其转化为SCAR(Sequence-characterized amplified region)标记。在156条10碱基随机引物中,引物S92(5′-CAGCTCACGA-3′)在小麦秆锈菌生理小种21C3CTH中扩增出一条782bp的特异片段,回收、克隆和测序该特异片段,根据特异片段的核苷酸序列设计了1对SCAR特异引物,经验证,该引物特异性良好。结果表明,小麦秆锈菌生理小种21C3CTH的RAPD标记被成功地转化为SCAR标记,这为该生理小种的分子鉴定和监测奠定了基础。     

利用小麦SSR标记追踪大赖草染色体Lr.2、Lr.7、Lr.14及其片段

定位于小麦7个部分同源群上的337对SSR引物中有113对SSR引物可检测到大赖草与普通小麦基因组间多态性，对小麦一大赖草Lr．2、Lr．7和Lr.14的异附加系和易位系的进一步分析表明，小麦7BL上的SSR引物gwm112在大赖草染色体Lr．2上具有特异扩增位点，可用来追踪Lr．2；5AS上的SSR引物gwm205、5AL的gwml56和5DL上的gwm212在Lr．7和Lr．14中均有特异扩增产物，可用于追踪Lr．7和Lr．14；而7AL上的SSR引物gwm63仅在大赖草染色体Lr．7上具有扩增位点。这不仅揭示了Lr．7可能存在第5和第7部分同源群染色体重排，而且也表明将引物gwm205、gwm156、gwm212与gwm63相结合可分别追踪大赖草Lr.7和Lr．14染色体及其片段。利用这些SSR引物可追踪同一植株中不同的大赖草染色体；与簇毛麦染色体6V上的SCAR标记相结合，能追踪同一植株中的大赖草和簇毛麦染色体片段。     

亚洲部分稻瘟病菌的交配型和雌性能育菌株的遗传多样性分析

 用稻瘟病菌两性菌株测定中国及亚洲部分国家稻瘟病菌有性时期的交配型和雌性能育菌株 ,并用SCAR分子标记对 14 3个菌株进行PCR扩增 ,分析它们的遗传多样性 ,得到亲缘关系树状图     

华山新麦草2Ns染色体SCAR标记的开发

为准确快速地建立小麦-华山新麦草杂交后代的分子标记鉴定方法,利用180条长度为10bp的随机引物R1～R180对小麦-华山新麦草全套(1Ns～7Ns)二体附加系及其亲本华山新麦草和普通小麦7182共9个材料进行了RAPD分析。结果表明,R131在华山新麦草和小麦-华山新麦草2Ns二体附加系中可以扩增出特异条带。将该特异条带回收并测序,发现其全长为1 126 bp,对其进行序列比对分析后设计SCAR引物S131,然后利用S131重新对9份材料进行了SCAR分析。结果显示,S131只在华山新麦草和小麦-华山新麦草2Ns二体附加系中扩增出特异性条带,表明RAPD标记R131已成功转化为可靠、特异的SCAR标记S131。这个新的SCAR标记可用于检测普通小麦背景下华山新麦草2Ns染色体。