Jain A, Ariyadasa R, Kumar A, Srivastava M N, Mohan M, Nair S
International Centre for Genetic Engineering and Biotechnology (ICGEB), Aruna Asaf Ali Marg, New Delhi, 110 067, India.
Theor Appl Genet. 2004 Nov;109(7):1377-84. doi: 10.1007/s00122-004-1774-x. Epub 2004 Aug 21.
Using amplified fragment length polymorphisms (AFLPs) and random amplified polymorphic DNAs (RAPDs), we have tagged and mapped Gm8, a gene conferring resistance to the rice gall midge (Orseolia oryzae), a major insect pest of rice, onto rice chromosome 8. Using AFLPs, two fragments, AR257 and AS168, were identified that were linked to the resistant and susceptible phenotypes, respectively. Another resistant phenotype-specific marker, AP19(587), was also identified using RAPDs. SCAR primers based on the sequence of the fragments AR257 and AS168 failed to reveal polymorphism between the resistant and the susceptible parents. However, PCR using primers based on the regions flanking AR257 revealed polymorphism that was phenotype-specific. In contrast, PCR carried out using primers flanking the susceptible phenotype-associated fragment AS168 produced a monomorphic fragment. Restriction digestion of these monomorphic fragments revealed polymorphism between the susceptible and resistant parents. Nucleotide BLAST searches revealed that the three fragments show strong homology to rice PAC and BAC clones that formed a contig representing the short arm of chromosome 8. PCR amplification using the above-mentioned primers on a larger population, derived from a cross between two indica rice varieties, Jhitpiti (resistant parent) and TN1 (susceptible parent), showed that there is a tight linkage between the markers and the Gm8 locus. These markers, therefore, have potential for use in marker-aided selection and pyramiding of Gm8 along with other previously tagged gall midge resistance genes [ Gm2, Gm4(t), and Gm7].
利用扩增片段长度多态性（AFLP）和随机扩增多态性DNA（RAPD）技术，我们已将赋予水稻抗稻瘿蚊（Orseolia oryzae）能力的基因Gm8定位到水稻第8号染色体上，稻瘿蚊是水稻的一种主要害虫。利用AFLP技术，鉴定出了两个分别与抗性和感病表型连锁的片段AR257和AS168。利用RAPD技术还鉴定出了另一个抗性表型特异性标记AP19(587)。基于片段AR257和AS168序列设计的SCAR引物未能揭示抗性亲本和感病亲本之间的多态性。然而，使用基于AR257侧翼区域的引物进行PCR显示出了表型特异性的多态性。相反，使用感病表型相关片段AS168侧翼引物进行的PCR产生了一个单态片段。对这些单态片段进行限制性酶切显示出了感病亲本和抗性亲本之间的多态性。核苷酸BLAST搜索表明，这三个片段与水稻PAC和BAC克隆具有很强的同源性，这些克隆形成了一个代表第8号染色体短臂的重叠群。使用上述引物对两个籼稻品种Jhitpiti（抗性亲本）和TN1（感病亲本）杂交产生的更大群体进行PCR扩增，结果表明这些标记与Gm8基因座之间存在紧密连锁。因此，这些标记有潜力用于标记辅助选择以及将Gm8与其他先前已标记的抗稻瘿蚊基因[Gm2、Gm4(t)和Gm7]进行聚合。
