Construction Of SSR Fingerprint Database And Core Collection Of Populus Clones

Poplar is the general name of Populus of Salicaceae.It has strong adaptability,fast growth,short rotation period,high wood yield and easy renewal and it is one of the fastgrowing timber species with the largest cultivation area in the middle latitude plain of the world.Under natural conditions,poplar is very easy to produce hybrids.The collected poplar germplasm is difficult to identify the differences by their morphological characteristics,and which is very easy to cause germplasm mixing.The molecular marker is a powerful tool for germplasm identification and core germplasm construction.In this study,based on TP-M13-SSR technology,the poplar clonal DNA fingerprint database and core collections had researched from the national poplar germplasm resource bank of Taigelin Paper Group Co.Ltd.The main research results are as follows:(1)Genetic diversity analysis of Germplasm14 pairs of SSR primers with clear bands and good repeatability were selected from 86 pairs of primers,141 poplar clones were amplified by SSR-PCR with these 14 pairs of primers,and 91 polymorphic bands were obtained.The invalid allele frequency of each SSR locus was calculated by Cervus software,and the primer ORPM3171 regarded as invalid allele was removed.The SSR amplification data of the remaining 13 pairs of primers were analyzed based on POPGENE software.The results showed that the number of alleles(NA)of 13 pairs of primers is 2～12,with an average of 6.8;The effective allele(NE)was 1.06～4.81,with an average of 2.60;Shannon’s information index(I)was 0.01～1.9,with an average value of 0.97;The polymorphism information content(PIC)was 0.19～0.81,with an average value of 0.56,and the observed heterozygosity(HO)was 0.06～0.76,with an average value of 0.40.Use NTSYS-pc2.1 software to analysis the 141 clones based on UPGMA method.These results showed that the clones were divided into three categories.(2)Construction of fingerprint databaseBased on the molecular marker primer combination method,two pairs of SSR primers were combined to evaluate the resolution of each pair of primers to the tested clones.Among the paired primers,GCPM1599 and GCPM1295 had the highest resolution,which could identify 53 clones,with a resolution of 37.85%;Among the three primer combinations,GCPM1599,GCPM1295 and PMGC2765 had the highest resolution,which could identify 113 germplasm,with a resolution of 80.71%;Among the four primer combinations,GCPM1599,GCPM1295,PMGC2765 and ORPM330 could identify 133 clones with a resolution of 95.00%;Among the five primer combinations,the combination of GCPM1599,GCPM1295,PMGC2765,ORPM330 and ORPM409 could identify 137 materials with a resolution of 97.85%;Among the 6 primer combination,the combination of GCPM1599,GCPM1295,PMGC2765,ORPM330,ORPM409 and ORPM214 can identify 140 materials(NL70 and NLS366 are the same genotype),and the resolution is 100%.In this study the unique fingerprint code belonging to each poplar clone germplasm material was constructed with these 6 pairs of primers.(3)Construction of core collectionEight primary core collections with sampling ratios of 10%,20%,30%,40%,50%,60%,70%and 80%were constructed by stepwise cluster random sampling method.Finally,30%was determined as the best sampling proportion,and 41 core collections were obtained.The number of alleles,observed heterozygosity,expected heterozygosity and Shannon’s information index of the core collection were 6.1538,0.5234,0.6378 and 1.3452 respectively,and the retention rates were 89.88%,104.84%,102.21%and 103.35%respectively.T-test showed that there was no significant difference in genetic diversity between 41 poplar clone core germplasm and the original germplasm;Principal coordinate analysis(PCoA)showed that the distribution was similar to that of the original germplasm,indicating that the core collection can better represent the original germplasm.This study shows that TP-M13-SSR technology can effectively identify poplar clones,clarify the genetic relationship between clones,and the results of genetic clustering are basically consistent with their pedigree relationship.The results of this study laid a foundation for the germplasm identification of poplar clones and the further development of poplar cross breeding.