Development of SCAR markers for early identification of cytoplasmic male sterility genotype in chili pepper (Capsicum annuum L.).
We previously used Southern blot analysis to detect restriction-length polymorphisms between male fertile and cytoplasmic male sterile (CMS) cytoplasms at the coxII and atp6 loci of the mtDNA of Capsicum annuum L. Two copies of atp6 were found in each male fertile and CMS pepper lines. Interestingly, one of the copies of atp6 in CMS pepper was a 3'-truncated pseudogene. The open reading frame of the coxII gene was the same in the fertile (N-) and CMS (S-) lines. However, the nucleotide sequence in the S-cytoplasm diverged from that in the N-cytoplasm 41 bp downstream of the stop codon. To develop CMS-specific sequence-characterized amplified region (SCAR) markers, inverse PCR was performed to characterize the nucleotide sequences of the 5' and 3' flanking regions of mitochondrial atp6 and coxII from the cytoplasms of male fertile (N-) and CMS (S-) pepper plants. Based on these data, two CMS-specific SCAR markers, 607 and 708 bp long, were developed to distinguish N-cytoplasm from S-cytoplasm by PCR. The CMS-specific PCR bands were verified for 20 cultivars containing either N- or S-cytoplasm. PCR amplification of CMS-specific mitochondrial nucleotide sequences will allow quick and reliable identification of the cytoplasmic types of individual plants at the seedling stage, and assessment of the purity of F1 seed lots. The strategy used in this report for identifying CMS-specific markers could be adopted for many other crops where CMS is used for F1 seed production.
Kim
Dong Hwan
DH
Department of Plant Science, College of Agriculture and Life Sciences, and Center for Plant Molecular Genetics and Breeding Research, Seoul National University, Seoul 151-921, Korea.
Kim
Byung-Dong
BD
D020413:3' Untranslated Regions / Q000235:genetics; D020121:5' Untranslated Regions / Q000235:genetics; D000251:Adenosine Triphosphatases / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D001483:Base Sequence; D002212:Capsicum / Q000235:genetics* / Q000378:metabolism; D051546:Cyclooxygenase 2 / Q000235:genetics* / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D004272:DNA, Mitochondrial / Q000235:genetics*; D018744:DNA, Plant / Q000032:analysis; D005819:Genetic Markers; D005838:Genotype; D007246:Infertility / Q000235:genetics*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D019105:Random Amplified Polymorphic DNA Technique; D017386:Sequence Homology, Amino Acid
D020413:3' Untranslated Regions; D020121:5' Untranslated Regions; D004272:DNA, Mitochondrial; D018744:DNA, Plant; D005819:Genetic Markers; D051546:Cyclooxygenase 2; D000251:Adenosine Triphosphatases