Development of EST-SSR and genomic-SSR in Chinese fir
In order to develop SSR primers of Cunninghamia lanceolata (Lamb.) Hook based on EST-SSR and genomic SSR, 292 non-repeated Chinese fir EST-sequences were assembled by removing low-quality and redundant fragments de-pend upon 444 ESTs sequences from NCBI Public database, and 143 genome-sequences were assembled by removing re-dundant fragments in 1 142 genome sequences. All those non-redundant sequences of Cunninghamia lanceolata (Lamb.) Hook was searched for mono-to hexa-nucleotide simple sequence repeats ( SSR or microsatellite) with software MISA, and the type,size and frequency of these SSRs were determined. Totally, There were 109 SSRs to be picked out among EST-sequences, the distribution density was 964.58 SSR/Mbp;and 39 SSRs were found in genome sequences, account-ing for the higher density of 1 037?24 SSR/Mbp. In both Genomic and EST sequence library, the Hexanucleotides re-peats, especially AT-rich repeat, was the most repeated type, and the AGC/CTG was the most trinucleotides repeats, The 37 and 95 pairs of SSR primers were designed based on EST library and Genomic by Primer 3.0, respectively. Ten EST-SSR primers and eight pairs of gSSR primers were selected randomly to perform PCR amplification with 12 individu-als. There were four pairs of primers showed polymorphism both in EST-SSR and gSSR, with polymorphic rate of 40%and 50%, respectively. Twenty-five polymorphic alleles were amplified by 8 SSR primer pairs, which showed an average 3?125 polymorphism alleles for each primer, the average effective alleles ( Na) 2.399 5, average PIC 0.519 1, average Hot 0?307 4. There were more polymorphism alleles from gSSR markers than EST-SSR among groups. More allelic loci were amplified with four pairs of primer from gSSR than the four pairs of primer from EST-SSR, and the former had high-er PIC value also.
国家科技期刊平台
NSTL
万方数据
维普
