Genetic mapping in Japanese coniferous tree species using microsatellite DNA markers

点击翻译按钮获取中文摘要

To establish an efficient selection system of SSR (simple sequence repeats) DNA regions, TAC (triplex affinity capture) that is a new method instead of a molecular hybridization method, a modified PIMA (PCR-based isolation of microsatellite arrays) and so on was investigated. The results were as follows:1.Using model DNAs in which [CT]-repeats ([CT]_<10>,[CT]_<15>,[CT]_<20>, and [CT]_<25>) were inserted, an optimization of TAC was executed on pH and temperature conditions of the reaction. In pH 6.4 -7.0, highly repeated DNA fragments were selected efficiently.2.The original PIMA method was modified for the reliable screening of the recombinant colony in which SSR was inserted.3.The modified TAC and PIMA were applied into actual screening of SSRs from the genomes of Cryptomeria japonica, Pinus thunbergii, Chamaecyparis obtuse, and Acacia mangium. In C.japonica, About 300 SSR fragments were selected, and SCAR (sequence characterized amplified region) markers were designed based on their sequences.4.The materials of families for constructing linkage-maps were made in C.japonica (a haploid-endospermic family of Sawara-1), P.thunbergii(a haploid-endospermic family of Sima-64), and C.obtuse (a cross-pollinated family between Aira-32 and Satsuma-8).