Development Of EST-SSR Primers And Backcross Population In Psathyrostachys Juncea

Russian wildrye（Psathyrostachys juncea）is a gramineous forage with excellent characteristics of cold resistance,drought resistance and light and medium salinity resistance.It has many uses such as forage production,soil and water conservation and environmental greening.However,since the cultivation of Russian wildrye in China is still in its infancy,the production and promotion of Russian wildrye in China are mostly introduced from the United States or Canada,and due to the relative lack of genetic background data of Russian wildrye,it has a certain impact on the utilization of germplasm resources and breeding improvement.Wild Russian wildryes germplasm in China has rich genetic background and is a high-quality hybrid breeding material.However,there is no detailed report on hybrid breeding and breeding process of Russian wildrye in China.This study reported the best combination process of Russian wildrye hybrid and explored the flowering habits of Russian wildrye.The best artificial pollination method were explored to reduce the impact of flowering time on seed yield and greatly improve the efficiency of hybrid seed production.In this study,Illumina Hiseq high-throughput sequencing platform was used to sequence the transcriptome of Russian wildrye,and SSR markers were developed according to the sequencing results.The screened EST-SSR characteristic primers of Russian wildrye were used for efficient and effective identification of F1 and BC1 hybrids.Finally,the heterosis performance of the two populations was studied by means of normal test,clustering and principal component analysis,and the improvement effect of agronomic traits of hybrid offspring was analyzed,which provided a basis for the selection of subsequent breeding materials.The main results showed below:（1）Using Illumina Hiseq high-throughput sequencing technology,a total of 38.81 Gb transcriptome data were obtained,a total of 100560 Unigenes were assembled,and 9300SSRs were identified in 29889 Unigenes above 1kb.The differentially expressed genes were screened according to the relative expression levels between the two samples,and a total of 2020 differentially expressed genes were identified,from which 400 Unigenes with SSRs loci were screened.EST-SSR with 10 tandem repeats was the most common,accounting for 24.75%of the total EST-SSR loci.The most abundant types are A/T,followed by GA/TC and GGC/GCC.（2）Among the 375 EST-SSR primers designed in this experiment,69 pairs of primers with similar annealing temperatures between the upper and lower primers were selected for amplification.Among them,54 primers could amplify the ideal product,of which 39 were polymorphic primers,and the amplification success rate was 78.26%.Sixteen pairs of polymorphic EST-SSR primers were randomly selected,among which three pairs of primers showed differences in the amplification products of F1 and BC1 parents and stable amplification in the hybrid offspring,which could be used for rapid identification of hybrid authenticity.The purity of F1 hybrids was 96.75%,and that of BC1 hybrids was 95.80%.The genetic similarity between the two populations was analyzed based on EST-SSR markers.It was found that the individual difference of backcross population was significantly less than that of F1 population,and the genotype was more similar,and more inclined to recurrent parent.（3）Flowering number is an important factor to determine the seed setting rate.The effects of meteorological factors such as temperature（X₁）,relative humidity（X₂）and ultraviolet index（X₃）on the flowering dynamics of Russian wildrye（Y）can be fitted and dataized,and the regression equation is Y=-103.57718+0.53508X₁+0.91837X₃.The flowering time and reproductive growth cycle of Russian wildrye can be predicted by combining the meteorological conditions and flowering habits of Russian wildrye.（4）The F1 generation population with abundant variation and obvious heterosis was obtained by crossing’XJ-ALT’as female parent and’Mengnong No.4’as male parent.71.67%of the fertile F1 plants showed excellent comprehensive agronomic traits and strong resistance,which could be selected for subsequent backcross materials according to the breeding objectives.The male parent’Mengnong No.4’was used as recurrent parent.A total of 102 BC1 Russian wildrye varieties were obtained,accounting for 74.45%of the total number of the BC1 population.Their comprehensive traits were mostly biased towards recurrent parent,which had the potential to continue trait improvement.