ISOLATION OF MICROSATELLITE MARKERS FROM APOSTICHOPUS JAPONICUS ESTs

Frequency and density of 2—6bp simple sequence repeats (SSRs) were analyzed from Apostichopus ja- ponicus EST (expressed sequence tag) database comprising 6632 sequences. The result shows that 416 microsatellite se- quences (taking 6.48%) were obtained. The dinucleotide repeats motifs appeared to be the most abundant type in A. ja- ponicus ESTs (taking 65.86%), followed by trinucleotide, tetranucleotide, pentanucleotide and hexanucleotide repeats at 26.68%, 1.68%, 0.24%, and 5.53%, respectively. 21 primer pairs were designed with Primer software and man- ual-searching method from EST sequences containing microsatellites. 13 primers had polymorphic PCR products in Chi- nese and Russian sea cucumber samples by electrophoresis analysis. In the 13 microsatellite loci, the alleles numbers dis- tributed from 3 to 8, and the range of these alleles were between 175 to 382bp. The observed heterozygosity and expected heterozygosity distributed from 0.158—0.650 and 0.198—0.841, respectively, which showed that these 13 microsatellite loci were suitable for genetic analysis.