In Silico Exploitation And Database Construction Of Microsatellite Markers In Tobacco(Nicotiana Tabacum L.)

Microsatellite,also termed as simple sequence repeat(SSR),widely exists in eukaryotic genomes.SSR has the features of rich variation,high polymorphism and codominance,and therefore is a kind of very useful molecular marker,which has been widely used in genetic mapping,gene mapping,germplasm identification,molecular marker-assisted breeding,and so on.Tobacco(Nicotiana tabacum L.)is an important economic crop.A great number of SSR markers have been developed in tobacco,and a high-density SSR genetic map of tobacco has been constructed.However,as the genetic diversity of tobacco is low,developing more SSR markers is necessary to meet the need of genetic research and molecular breeding in tobacco.With the rapid development of the next-generation sequencing technology,the genomes of three tobacco varieties(BX,TN90 and K326)have been sequenced and published,providing valuable data for large-scale exploitation of tobacco SSR markers.In this study,using these data,tobacco SSRs were completely searched and analyzed,primers were designed and polymorphisms were tested to develop SSR markers,and part of the newly developed SSR markers were mapped onto the tobacco genetic map according to their linkage relationship with the published SSR markers.In addition,a tobacco SSR database was constructed for the results obtained.The main results are as follows:(1)A total of 630,742 SSRs were found,of which 14.44%were common in three varieties,27.97%were common in two varieties,and 57.58%were specific in single varieties.(2)Among different SSR types,two-base repeats had the largest number,followed by three-base repeats,and the third was five-base repeats.(3)In terms of the motifs,A/T was more than G/C in the single-base motifs;AT/TA was the most and GC/CG was the fewest among the two-base motifs;and AAC/GTT were the most and CGC/GCG were the fewest among the three-base motifs.In general,the higher in GC content,the lower in number.(4)In the three varieties,about 3/5 SSRs had PCR primers successfully designed,having the potential to be exploited as molecular markers.(5)Among the SSRs common in two or three varieties,17.39%displayed polymorphisms among the varieties,of which the two-base repeats were the most,followed by the three-base repeats.(6)The published tobacco SSRs were mapped onto the scaffolds of the three varieties via e-PCR.This allowed the scaffolds to be mapped onto the tobacco genetic map.Furthermore,according to the position relationship on the scaffolds,a great many(?46,000)newly developed SSR markers were mapped onto the genetic map,making the marker density of the genetic map increased more than 20 times.The results of this study greatly increased the number of tobacco SSR markers,which will enhance the development of genetic studies and molecular breeding in tobacco.