Identification of gentian cultivars using SCAR markers based on intron-length polymorphisms of flavonoid biosynthetic genes

Gentian is one of the most important ornamental flowers in Japan. Gentiana triflora, G. scabra and their interspecific hybrids are the main breeding materials. Gentian cultivars are easily proliferated vegetatively, therefore it is important to develop a reliable discrimination method to prevent the illegal propagation and distribution of various high-value cultivars. Here, we report five sequence characterized amplified region (SCAR) markers based on the length polymorphisms in introns of four gentian flavonoid biosynthetic genes. These SCAR markers effectively discriminated nine gentian cultivars and nine breeding lines. This method could be applied in identifying gentian cultivars/lines and therefore will aid in protecting breeders’ rights.     

鉴定与芥菜型油菜紫叶基因Bjpl1紧密连锁的AFLP及SCAR标记

叶色性状,作为一个形态学标记,在作物的遗传改良中扮演着重要的角色。在芥菜型油菜品系1280中发现了一个叶色自发突变体,将其命名为1280 1。1280 1新叶通常呈绿色,叶缘和叶脉紫色。随着叶片生长,整个叶片均呈现紫色。1280 1目前已自交7代,叶片紫色性状已稳定。为进一步研究1280 1的遗传特性,将1280 1与2个芥菜型油菜绿叶亲本（芥菜型多室油菜和富源油菜）进行杂交, F₁自交后获得F₂群体,同时F₁与芥菜型油菜绿叶亲本进行回交得到BC₁分离群体。F₂分离群体统计表明,1280 1的紫叶性状受到1对显性基因控制,将该基因命名为 Bjpl1;利用AFLP结合BSA的方法,设计512对引物组合,在1280 1与芥菜型多室油菜构建的BC₁分离群体筛选与Bjpl1基因紧密连锁的分子标记,共获得10对与目标基因紧密连锁的分子标记。其中,PL02、PL07 和 PL10与Bjpl1基因共分离;PL07被转化成一个SCAR标记。目标基因两侧最近的标记分别为PL01 (PL04) 和 PL08,它们与目标基因间的遗传图距分别为0.7和1.3 cM。     

南方、爪哇和花生根结线虫的快速灵敏的PCR鉴定方法

 为了研制南方、爪哇和花生根结线虫快速灵敏的检测和鉴定方法,分别分离了4个南方根结线虫和3个爪哇根结线虫特异性的随机扩增多态性DNA (RAPD)片段。在这些RAPD标记DNA序列的基础上,设计了多对SCAR PCR引物,并用源于国内外的南方、爪哇、花生、北方和象耳豆根结线虫群体验证其扩增特异性和灵敏度。最终确定了3对高效扩增的SCAR引物,它们组合使用可以可靠灵敏地鉴定南方、爪哇和花生根结线虫。3对引物的扩增灵敏度达1/3条的二龄幼虫、雄虫或雌虫,这表明本研究研制的PCR鉴定法可用于生产实践中土样和根样中3种根结线虫快速灵敏的鉴定。     

稻曲病菌的SCAR标记及其PCR检测

为建立稻曲病菌Ustilaginoidea virens快速、灵敏的PCR早期检测技术,以S380为引物,对稻曲病菌和其它参试病原菌的DNA进行RAPD-PCR扩增。稻曲病菌RAPD特异片段经回收、克隆和测序后,根据序列用Primer 5.0软件设计了特异性SCAR引物US-SF（5’-TCGCCTCAGACCTCAATC-3’）/US-SR（5’-GAGCCTCAAATGCCTTCC-3’）和巢式PCR引物US-NF（5’-AGCGTCTCCTGCAACC-AC-3’）/US-NR（5’-GAGCCTCAAATGCCTTCC-3’）。利用引物US-SF/US-SR通过常规PCR对供试稻曲病菌均可扩增出1条大小约257 bp的清晰条带,对其它植物病原菌DNA的PCR产物均无扩增条带,最低可检测到1 pg/μL 的病菌基因组DNA;而利用引物US-SF/US-SR和US-NF/US-NR通过巢式PCR可从10 fg/μL的病菌基因组DNA中扩增出1条大小约210 bp的特异性条带。试验表明,巢式PCR检测灵敏度比常规PCR提高了100倍,且巢式PCR可从接菌的水稻幼颖和自然感染的谷粒中检测出稻曲病菌。     

一个鉴别低温刺激型秀珍菇的ISSR-SCAR标记建立与应用

为了快速、准确地鉴别低温刺激型秀珍菇菌株,在对18个供试菌株进行拮抗试验、现蕾出菇试验、ISSR图谱分析的基础上获得了秀珍菇菌株的ISSR特异性标记,将其克隆、测序.根据测序结果,应用Primer 6.0软件设计SCAR引物进行引物筛选,获得1对引物并成功转化为低温刺激型秀珍菇的稳定SCAR标记.采用该引物对供试秀珍菇...     

Identification of Molecular Markers Linked to TuMV Resistant Gene in Cabbage

A total of 144 F2 individuals were obtained from the crossing between 1047 （susceptible） and A21 （resistant）. Two RAPD markers were screened out in 200 random primers using：BSA（Bulked Segregant Analysis）. Two RAPD markers, designated as AG13/2000 and U16/660, were 7.7 cM and 8.38 cM apart from the TuMVresistant gene, respectively. The two RAPD fragments were converted to SCAR markers. SCAR markers were confined in germplasm.     

普通小麦-奥地利黑麦抗条锈病衍生系NR1121的鉴定

【目的】检测从普通小麦(Triticum aestivumL.)与奥地利黑麦杂交后代选育出的、形态学稳定的抗条锈病衍生系NR1121的黑麦遗传物质。【方法】在室内条锈病抗性鉴定的基础上,采用细胞学方法、基因组原位杂交、SCAR(Sequence characterized amplified region)标记以及SSR(Simple sequence repeat)标记、A-PAGE等方法,对普通小麦-奥地利黑麦衍生系NR1121进行分子细胞遗传学鉴定。【结果】NR1121和奥地利黑麦对条中32号生理小种(CY32)免疫,陕麦611和携带Yr9基因的洛夫林10、洛夫林13、秦麦9号、丰抗8号、陕229和偃师9号均高感,表明NR1121携带的抗条锈病基因来源于奥地利黑麦。NR1121细胞学稳定,根尖细胞染色体数2n=42,花粉母细胞减数分裂期2n=21Ⅱ;以奥地利黑麦总基因组DNA为探针的原位杂交结果显示,NR1121含有2条奥地利黑麦染色体。SCAR和SSR标记(Xgwm 131-1B)检测结果表明,NR1121携带黑麦1R染色体的遗传物质。A-PAGE结果显示,NR1121在ω区的Gli-B1位点具有黑麦特征带,说明NR1121含有黑麦1R染色体短臂上的遗传物质。【结论】NR1121为农艺性状优良的小麦-黑麦1R异代换系,对CY32免疫,其抗病基因来源于奥地利黑麦,可能是一个不同于Yr9基因的新抗条锈病基因。该种质可作为条锈病抗源用于小麦抗病育种。     

利用SSR标记技术构建黄瓜遗传连锁图谱

利用抗黄瓜花叶病毒病(CMV)的黄瓜材料F-3和感CMV的黄瓜材料HZL04-1为亲本,构建包含190个F2后代的遗传作图群体。结果表明:采用SSR、EST-SSR和SCAR 3种分子标记技术对该群体进行遗传连锁分析,获得1个包含170个SSR标记,6个EST-SSR标记,3个SCAR标记的黄瓜遗传连锁图谱,该图谱涉及7个连锁群,总长度为862.886 cM,平均图距为4.79 cM。该图谱的构建为黄瓜抗CMV的QTL定位奠定了基础,并确定了连锁群与染色体之间的对应关系。     

辣椒恢复基因SSR标记定位及分子标记辅助选择育种

利用175对均匀分布在辣椒染色体上的SSR引物对辣椒不育系113A、恢复系139及其F2代群体进行筛选,发现AF208834引物和Rf基因连锁,遗传距离为20.8cM。用CRF-SCAR标记对36份恢复系材料进行分子标记辅助选择验证,结果表明,30份恢复系有特异条带,正确率为83.33%,并从108份未知育性材料中,检测到41份有恢复基因的材料;同时将SSR标记和SCAR标记结合起来,建立了与育性基因有关的多重PCR技术,进行辣椒恢复基因及其等位基因纯合状态的选择,显著提高了辣椒雄性不育三系的选择效率。     

Analysis on ‘Whole Red’ of patterns Oujiang colour carp (Cyprinus carpio var.color ) by SRAP and SCAR markers

The sequence‐related amplified polymorphism (SRAP) technique was applied to identify the genetic difference between the ‘Whole Red’ (‘WR’) population and the ‘Whole White’ (‘WW’) population. The specific SRAP band, namely SR_2,7^{173 bp} (SR indicates the first two letters of SRAP) from ‘WR’ and ‘WW’, was identified from the amplified bands of 12 primer pairs (PP) screened from 88 SRAP PP. After gel extraction, cloning and sequencing of specific band from the two populations, the sequence was submitted to database of Genome Sequence Survey. blast analysis showed that this SRAP fragment shared high similarity to functional genes of Danio rerio. Four primers (22–26 bases) were designed according to the sequence information. Then, polymerase chain reaction amplification was carried out in the two populations. The experimental results also showed that among four sequence‐characterized amplified region (SCAR) markers, only SC‐3 (154 bp) developing from SR_2,7^{173 bp} (me2‐em7) showed specificity to ‘WR’. Examination with a large sample size showed that SCAR 3 (SC‐3) could be positively amplified with 85% frequency in the ‘WR’ population rather than in the ‘WW’ population. The results indicated that the SC‐3 marker could be used as a specific molecular marker for population identification, providing an effective, easy and rapid method for discriminating different populations and conducting genetic analysis.