Promoter anchored amplified polymorphism based on random amplified polymorphic DNA (PAAP-RAPD) in cotton

Mingxiong Pang; Percy, R. G.; Hughs, Ed.; Jinfa Zhang
June 2009
Euphytica;Jun2009, Vol. 167 Issue 3, p281
Academic Journal
Non-coding sequences account for a majority of the higher plant genome, some of which have important effects in gene regulation and plant development. In an effort to develop molecular marker systems to search for polymorphisms associated with high fiber yield and quality in cotton, we have developed a methodology that could specifically target the regulatory regions of the cotton genome. In this study we designed 10-nucleotide degenerate promoter primers based on conserved core promoter sequences and tested their applicability in PCR amplifications in combination with 10-mer random amplified polymorphic DNA (RAPD) primers. The amplified markers are called promoter anchored amplified polymorphism based on RAPD (PAAP-RAPD). Forty cotton genotypes with diverse genetic and geographical backgrounds were used to test the PAAP-RAPD system using polyacrylamide gel electrophoresis. Based on PAAP-RAPD markers amplified from 12 primer combinations, the 40 genotypes were classified into five distinctive groups: two Upland cotton ( Gossypium hirsutum) groups from China, another two Upland cotton groups from the USA, and one group from American Pima cotton ( G. barbadense). The groupings are in general consistent with their genetic and geographical origins. Thirty-six PAAP-RAPD and RAPD fragments were cloned and four of them were further subjected to sequence analysis. Signal scanning using software PLACE confirmed that they contained an array of cis-regulatory sequences in addition to the core promoter sequences. The results demonstrate the potential application of PAAP-RAPD as a new marker system specifically targeting regulatory regions of the plant genome.


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