Use of capillary array electrophoresis single-strand conformational polymorphism analysis to estimate genetic diversity of candidate genes in germplasm collections.
Capillary array electrophoresis single-strand conformation polymorphism (CAE-SSCP) analysis provides a reliable high-throughput method to genotype plant germplasm collections. Primers designed for highly conserved regions of candidate genes can be used to amplify DNA from plants in the collection. These amplified DNA fragments of identical length are turned into useful markers by assaying sequence differences by CAE-SSCP analysis. Sequence differences affect the electrophoretic mobility of single-stranded DNA under non-denaturing conditions. By collecting the mobility data for both strands assayed at two temperatures, alleles can be defined by mobility alone. For a germplasm collection with an unknown number of alleles at a locus, such mobility data of homozygotes can be used to determine the number of unique alleles without the necessity of cloning and sequencing each allele.
Kuhn
David N
DN
Department of Biological Sciences, Florida International University, Miami, Florida 33199, USA.
Schnell
Raymond J
RJ
D000483:Alleles; D002021:Buffers; D017931:DNA Primers; D018744:DNA, Plant / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D003627:Data Interpretation, Statistical; D019075:Electrophoresis, Capillary / Q000379:methods* / Q000592:standards; D020224:Expressed Sequence Tags; D005456:Fluorescent Dyes; D017343:Genes, Plant; D005821:Genetic Techniques / Q000592:standards / Q000706:statistics & numerical data; D014644:Genetic Variation; D008970:Molecular Weight; D010944:Plants / Q000235:genetics; D011108:Polymers; D018807:Polymorphism, Single-Stranded Conformational; D013696:Temperature
D002021:Buffers; D017931:DNA Primers; D018744:DNA, Plant; D005456:Fluorescent Dyes; D011108:Polymers