Development of reliable PCR-based markers linked to downy mildew resistance genes in lettuce

Abstract

Sequence characterized amplified regions (SCARs) were derived from eight random amplified polymorphic DNA (RAPD) markers linked to disease resistance genes in lettuce. SCARs are PCR-based markers that represent single, genetically defined loci that are identified by PCR amplification of genomic DNA with pairs of specific oligonucleotide primers; they may contain high-copy, dispersed genomic sequences within the amplified region. Amplified RAPD products were cloned and sequenced. The sequence was used to design 24-mer oligonucleotide primers for each end. All pairs of SCAR primers resulted in the amplification of single major bands the same size as the RAPD fragment cloned. Polymorphism was either retained as the presence or absence of amplification of the band or appeared as length polymorphisms that converted dominant RAPD loci into codominant SCAR markers. This study provided information on the molecular basis of RAPD markers. The amplified fragment contained no obvious repeated sequences beyond the primer sequence. Five out of eight pairs of SCAR primers amplified an alternate allele from both parents of the mapping population; therefore, the original RAPD polymorphism was likely due to mismatch at the primer sites.

Molecular Mechanisms of Epigenetic Variation in Plants

“…Additional marker strategies were developed using different combinations of PCR, restriction digestion and gel electrophoresis techniques. Major marker techniques included random amplification of polymorphic DNAs (RAPDs) [4]; sequence characterized amplified region (SCARs) [5]; cleaved amplified polymorphic sequences (CAPS) [6]; Intersimple Sequence Repeats (ISSRs) [7]; amplified fragment length polymorphisms (AFLPs) [8]; and direct amplification of length polymorphism (DALP) [9]. The improvement of Sanger sequencing throughput in the in combination with the start of genome and expressed sequence tag (EST) sequencing programs in model plant species, led to the acceleration in the identification of variation at the single base pair resolution [10].…”

Genetic Improvement of Maize by Marker-Assisted Breeding

“…이처 럼 식물체 유전체에서는 빈번한 SNP 다형성 때문에 mapping, MAS 그리고 유전자 지도 기반 클로닝(map-based cloning) 이 가능하게 되었다 (Batley et al, 2003 (Olson et al, 1989). 반면에 SCAR은 STS 의 장점을 공유하면서도 2가지 측면에서 차이가 있는데, 첫 째는 SCAR는 우선적으로 유전적으로 정의되어 있기 때문 에(primarily defined genetically) 유전체에서 물리적 표시 인자(physical landmarker) 뿐만 아니라 유전적 마커로서의 기능도 할 수 있으며, 둘째는 SCAR은 오직 PCR에 의해서 만 분석되기 때문에 반복되는 구간을 가지고 있다는 특징이 있다 (Paran & Michelmore, 1993).…”

Development of RAPD-SCAR Molecular Marker Related to Seed-hair Characteristic in Carrot

“…그러나 RAPD 방법은 10개 정도의 nucleotide의 짧은 random primer를 이용하여 낮은 annealing 온도에서 증폭이 일어나기 때문에 반응조건 에 따라 결과가 달라질 수 있다 (Cutler et al, 2006 (Negi et al, 2000;Paran and Michelmore, 1993).…”