Singh Monika, Chaudhary K, Boora K S
Department of Biotechnology and Molecular Biology, CCS Haryana Agricultural University, Hisar, 125 004, Haryana, India.
Theor Appl Genet. 2006 Dec;114(1):187-92. doi: 10.1007/s00122-006-0423-y. Epub 2006 Oct 25.
Anthracnose, caused by Colletotrichum graminicola, infects all aerial parts of sorghum, Sorghum bicolor (L.) Moench, plants and causes loss of as much as 70%. F(1) and F(2) plants inoculated with local isolates of C. graminicola indicated that resistance to anthracnose in sorghum accession G 73 segregated as a recessive trait in a cross with susceptible cultivar HC 136. To facilitate the use of marker-assisted selection in sorghum breeding programs, a PCR-based specific sequence characterized amplified region (SCAR) marker was developed. A total of 29 resistant and 20 susceptible recombinant inbred lines (RILs) derived from a HC 136 x G 73 cross was used for bulked segregant analysis to identify a RAPD marker closely linked to a gene for resistance to anthracnose. The polymorphism between the parents HC 136 and G 73 was evaluated using 84 random sequence decamer primers. Among these, only 24 primers generated polymorphism. On bulked segregant analysis, primer OPA 12 amplified a unique band of 383 bp only in the resistant parent G 73 and resistant bulk. Segregation analysis of individual RILs showed the marker OPA 12(383) was 6.03 cM from the locus governing resistance to anthracnose. The marker OPA 12(383) was cloned and sequenced. Based on the sequence of cloned RAPD product, a pair of SCAR markers SCA 12-1 and SCA 12-2 was designed using the MacVector program, which specifically amplified this RAPD fragment in resistant parent G 73, resistant bulk and respective RILs. Therefore, it was confirmed that SCAR marker SCA 12 is at the same locus as RAPD marker OPA 12(383) and hence, is linked to the gene for resistance to anthracnose.
由禾生炭疽菌引起的炭疽病会感染高粱（双色高粱）植株的所有地上部分，造成高达70%的损失。用当地分离的禾生炭疽菌接种F(1)和F(2)植株，结果表明高粱种质G 73对炭疽病的抗性在与感病品种HC 136的杂交中表现为隐性性状分离。为便于在高粱育种计划中使用标记辅助选择，开发了一种基于PCR的特异性序列特征扩增区域（SCAR）标记。从HC 136×G 73杂交后代中选取了29个抗性和20个感病重组自交系（RILs）用于混合分离群体分析，以鉴定与炭疽病抗性基因紧密连锁的RAPD标记。使用84个随机序列十聚体引物评估亲本HC 136和G 73之间的多态性。其中，只有24个引物产生了多态性。在混合分离群体分析中，引物OPA 12仅在抗性亲本G 73和抗性混合群体中扩增出一条383 bp的独特条带。对单个RILs的分离分析表明，标记OPA 12(383)与控制炭疽病抗性的基因座相距6.03 cM。标记OPA 12(383)被克隆并测序。基于克隆的RAPD产物序列，使用MacVector程序设计了一对SCAR标记SCA 12-1和SCA 12-2，它们在抗性亲本G 73、抗性混合群体和各自的RILs中特异性扩增该RAPD片段。因此，证实SCAR标记SCA 12与RAPD标记OPA 12(383)位于同一基因座，因此与炭疽病抗性基因连锁。
