Cot-based sampling of genomes for polymorphic low-copy DNA
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Cot-based sampling of genomes for polymorphic low-copy DNA Junkang Rong • Jon S. Robertson • Stefan R. Schulze • Andrew H. Paterson
Received: 22 March 2010 / Accepted: 9 September 2010 / Published online: 1 October 2013 Ó Springer Science+Business Media Dordrecht 2013
Abstract DNA polymorphisms are powerful tools for many evolutionary and genomic studies in plants including molecular breeding. Single nucleotide polymorphisms (SNPs) are the most elemental DNA marker for genomic studies, but even with advances in DNA sequencing technology, SNP discovery remains costly and computationally demanding, especially in large genomes that are rich in repetitive DNA such as those of many plants. Here we report a method using DNA renaturation kinetics (Cot techniques), sequencing, and BLAST-based screening to identify low-copy, non-coding DNA sequences that were subsequently found to be relatively rich in polymorphisms. A total of of 63 such fragments isolated from a diploid D genome cotton species (Gossypium raimondii) revealed a higher frequency of polymorphisms than that observed for cotton expressed sequence tags or hypomethylated (PstI-susceptible) genomic DNA. While microsatellite-derived loci show still higher
J. Rong J. S. Robertson S. R. Schulze A. H. Paterson (&) Plant Genome Mapping Laboratory, University of Georgia, Athens, GA 30602, USA e-mail: [email protected] J. Rong e-mail: [email protected] J. Rong School of Agriculture and Food Science, Zhejiang A and F University, Linan, 311300 Zhejiang, People’s Republic of China
polymorphism rates, they often fall in repetitive elements and their sequence analysis is often complicated by alignment difficulties. The potential applications of Cot-filtered noncoding (CFNC) DNA in development of DNA markers are discussed. Keywords
Cotton SNP Cot DNA Marker
DNA polymorphisms in plants are powerful tools for many evolutionary and genomic studies including molecular breeding. For example, DNA polymorphism can be used to (1) determine the parental origins of chromosome segments across the genome in progenies of crosses between different genotypes, (2) map traits of interest as a foundation for markerassisted selection, and (3) clone genes associated with the polymorphism. DNA polymorphism can also be used to study species evolution and genomic structures of varieties and species. DNA polymorphisms can be classified into two types: base substitutions (involving single nucleotides or extending over several nucleotides) and insertions/deletions (also single nucleotide or extended). Low-copy DNA is a preferred source of polymorphisms because of increased confidence that orthologous (corresponding) loci are being compared in different individuals. Most genes occur in single or low-copy numbers; however, genes (in particular the coding regions of genes) tend to be functionally constrained and therefore tolerate only a subset of the possible range of DNA polymorphisms.
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In addition to genes, it has long been known that higher plant gen
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