Genetic Analysis Of Pomegranate In China Based On EST-SSR Markers

Pomegranate(Punica granatum L.)is a high edible,medicinal and ornamental value fruit tree which belongs to Punicaceae,Punica L.Through the construction of cDNA Library of pomegranate and development of EST-SSR primers,38 pomegranate accessions collected were analyzed.These research provide reference in identification of varieties,Hybridization breeding and Molecular mark assisted breeding for Chinese pomegranate.The following is the result:1.Using the mature seed of soft-seed pomegranate cultivar Hongmanaozi as materal,pomegranate cDNA libraries were constructed.Then a set of clones were randomly selected and single-pass sequenced from their 5?end.As a result,1596 valid sequences were generated,in which 907 unigenes were obtained after initial assembly with Clustalx program,179 bitype to tetra-type SSRs(19.7%)were obtained from these unigenes.Of the EST-SSRs,dinucleotide and trinucleotide have a abosolute majority,the most common two repeat motifs were GA(24.0%)and TG(21.2%)in dinucleotide,and the most common three repeat motifs in trinucleotide are closed: AGA(10.4%),AAG(9.0%),GAG(9.0%).2.Twenty primer pairs were design by Primer5.0 under the conditions of setting.And 10 primer pairs were as the same as expected,using screening means by three cultivas(baiyushizi,taishanhong and pishan no.2)from different areas.3.Ten primers were screened by polyacrylamide gel electrophoresis.In the end,8 primers with high polymorphism were screened successfully,detecting 2.60 alleles(mean).The observed heterozygosity(Ho)and polymorphic information content(PIC)values ranged from 0.125 to 0.833(mean = 0.367)and from 0.080 to 0.664(mean = 0.377),respectively.4.Genetic diversity of the accession set and cluster analysis was elucidated using the newly-developed markers,revealing a distinctive genetic background of Chinese pomegranate,The similarity coefficient values ranged from 0.68 to 0.97 with a mean of 0.83.All the accessions were clustered into two major groups(I and II)at a similarity level of 0.68 by the dendrogram.Most of Anhui accessions were clustered in group I and most Shandong accessions in group II,revealing a different genetic background between two groups.Group I were further divided in 3 subgroups(I-A,I-B,I-C)at a similarity level of 0.80,and all the accessions in subgroup I-B come from Huaibei,anhui area,proving a unique genetic background in Huaibei.