Development Of EST-SSR And Evaluation Of Genetic Diversity Of Common Millet(Panicum Miliaceum)

Common millet(Panicum miliaceum L.)is an ancient small crop that is drought tolerant,resistant to infertility and nutritious,and is of great significance for sustainable ecological agriculture and crop ecological diversity.Because of the complex heterotetraploid structure and evolutionary conservation of common millet,We know little about its genomic function,and the genetic diversity of common millet has not been fully evaluated.With the advent of the second generation of high-throughput sequencing technology,a large number of SSR(Simple Sequence Repeat)markers can be developed to study the genetic diversity of the common millet population,aiming at germplasm identification,specific gene mining,germplasm conservation and species geographic origin.The kinship research provides a theoretical basis.This study was based on high-throughput sequencing to develop polymorphic EST-SSR markers,and used markers to assess the genetic diversity of 144 domestic and foreign common millet resources.Below are key research findings:(1)Development of EST-SSR markers for common millet200 pairs of primers were designed from 1000 SSR sequences and polymorphism screening by 6common millet materials.The 23 pairs of primers could not amplify DNA bands,177 pairs(88.5%)had amplified bands,and the bands were blurred or present.The 80 pairs of primers were polymorphic and the development efficiency was 40%.(2)Evaluation of 80 markersThe 80 EST-SSR markers obtained,10 single-base repeats(50%)were mainly A/T repeats(50%each);15 two-base repeats,mainly AG repeats(40%);55 three-base repeats(38.19%),the main types are GGC,GCG,GCC(13%,11%,9%).The EST-SSR primers ranged in size from 50-500 bp,the RYW140 allele band was the most(20 sites),and the RYW134 allele band was the least(5 sites).Primer resolution(Rp)values were 0.67-4.67(mean 2.00),RP values were 0.33-0.83(average 0.55).The Rp values of 80 markers were generally distributed in the range of 1-2.Within the Rp values range of 4-5,the number of markers was the least and the marks with higher Rp values were mainly RYW107,RYW161,RYW166,RYW115 and RYW158.The main genetic polymorphisms in the markers are RYW150,RYW168,RYW124,RYW137,RYW110,RYW160,RYW163,RYW114,RYW125 and RYW158,and the markers with lower genetic diversity are mainly RYW103,RYW109,RYW167 and RYW144.The genetic parameters of the two-base repeat marker(Shannon's information inddex(I)is 0.8339)were similar to the three-base repeat marker(I is 0.8312),and the single base repeat marker was the lowest(I is 0.7497).(3)Genetic diversity analysis of 144 common millet resourcesA total of 206 allelic variations were detected in 80 polymorphic loci.The average value of I was0.8599,the average PIC value was 0.4573,the genetic parameter of RYW124 was the highest,and the RYW108 was the lowest.The richness of genetic diversity in each region is Northern spring-sowing ecotope(NSP)> Loess Plateau spring and summer-sowing ecotope(LPSS)> Northern summer-sowing ecotope(NSU)> Northeast spring-sowing ecotope(NES)> Northwest spring and summer-sowing ecotope(NWSS)> Southern autumn and winter-sowing ecotope(SAW).The genetic distance between the NSP and LPSS was the smallest(0.0117),and the genetic agreement was the highest(0.9884).Landrace > Abroad >Bred variety > Wild germplasm,Overall,Domestic > Abroad.Based on UPGMA,144 medlars were grouped into groups A,B and C.The first two groups were mainly materials of the spring scorpion in the north,and the group C was mainly the material of the spring and summer scorpion in the Loess Plateau.Group C can be divided into two subgroups,C1 and C2,mainly for the northern spring scorpion area and foreign resource materials,with the lowest genetic diversity parameters.The C1 subgroup can be divided into two sub-Asian groups,C11 and C12,which are mainly the northern spring scorpion area and foreign resource materials.The genetic relationship between resource groups in different regions is basically related to their geographical distribution.Structure is used to divide the test materials into 2 and 4 groups.When K=2,it is divided into red and green groups,mainly for foreign and northern resource gene pools.When K=4,it is divided into 4 groups.The red and green groups are mainly the northern resource gene pool,and the red group has the highest genetic diversity parameters.The blue and yellow groups are mainly the Loess Plateau and foreign gene banks.More than half of the red group materials with K=2 are not genetically differentiated,that is,the yellow group with K=4,representing the foreign group gene pool.The genetic group of foreign gene banks is the lowest,and there are obvious genetic differentiation between domestic and foreign resources.Through principal component analysis(PCA),PC1,PC2,and PC3 accounted for 22.9%,2.9%,and2.2% of the total gene variation,respectively,with a cumulative variation of 28%.Based on the SSR marker genotype,PC1 divided 144 resources into 7 different groups,and the results were consistent with the geographic origin of the material.