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Development of SCAR marker linked to a root-knot nematode resistant gene in peanut

机译：与花生根结线虫抗性基因相关的SCAR标记的开发

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摘要

Root-knot disease caused by Meloidogyne spp. is the most important nematode disease of peanut. Even though many management strategies have been applied to control this disease on peanut, resistance is the most recommendable. Marker-assisted selection has been used as a useful tool for screening of resistant individuals in segregating populations. However, it requires many laborious steps. Thus, there is a need for PCR - based markers, which are more practical, rapid, and efficient. In this study, we tried to develop a SCAR marker linked to root-knot nematode resistance locus in peanut based on the RFLP marker R2430E. The entire sequence of R2430E was 2217 bp and contained one putative open reading frame (ORF) of 713 nucleotides. Thirteen primers including 5 forward and 8 reverse primers were synthesized to sequence the entireR2430E. Based on the results of BLAST searches, R2430E appeared to encode an AAA ATPase containing von Willebrand factor type A (VWA) domain from Magnetococcus sp. MC-1 (106 bits). To determine if there is a portion of the R2430E that hybridizes only to a band co-segregating with the resistance locus, we generated 4 probes spanning different parts of the gene. Southern analysis using these probes revealed identical banding patterns for each probe. Therefore, we concluded that there is very limited if any sequence polymorphism between different alleles detected by the R2430E probe. Additionally, this conclusion is supported by the experiment in which we tested 25 primer pairs derived from the R2430E using genomic DNA from both resistance and susceptible genotypes. In this experiment, all primer pairs amplified identical PCR fragments, suggesting again that there is little or no sequence divergence between putative alleles as differentiated by southern blotting. To identify possible single nucleotide polymorphisms (SNPs) between polymorphic R2430E RFLP bands, we cloned several fragments that span the entire R2430E transcribed sequence. Surprisingly, no SNPs were identified in the transcribed region of this gene. We propose that polymorphism detected by this RFLP marker is outside of the R2430E.
机译：根结线虫引起的根结病。是花生最重要的线虫病。尽管已采用许多管理策略来控制花生上的该病，但最可取的是抗药性。标记辅助选择已用作筛选隔离人群中抗性个体的有用工具。但是，这需要许多费力的步骤。因此，需要更实用，快速和有效的基于PCR的标记。在这项研究中，我们试图基于RFLP标记R2430E开发与花生根结线虫抗性基因座相关的SCAR标记。 R2430E的整个序列为2217 bp，包含一个713个核苷酸的推定开放阅读框（ORF）。合成了包括5个正向和8个反向引物的13个引物来对整个R2430E进行测序。根据BLAST搜索的结果，R2430E似乎编码了一个来自磁球菌属物种的von Willebrand因子A型（VWA）域的AAA ATPase。 MC-1（106位）。为了确定R2430E是否存在仅与与抗性基因座共分离的条带杂交的部分，我们生成了4个跨越基因不同部分的探针。使用这些探针的Southern分析显示每种探针具有相同的条带模式。因此，我们得出的结论是，R2430E探针检测到的不同等位基因之间的序列多态性是否非常有限。此外，该结论得到了实验的支持，在实验中，我们使用了来自抗性和易感基因型的基因组DNA测试了来自R2430E的25对引物。在该实验中，所有引物对均扩增了相同的PCR片段，这再次表明，通过Southern印迹法区分的推定等位基因之间几乎没有序列差异。为了确定多态性R2430E RFLP条带之间可能的单核苷酸多态性（SNP），我们克隆了跨越整个R2430E转录序列的几个片段。出乎意料的是，在该基因的转录区中未鉴定出SNP。我们建议该RFLP标记检测到的多态性不在R2430E内。

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    Yang Hee Jeong;

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  • 年度 2004
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