Abstract:

Due to the narrowing of genetic differences between alfalfa （Medicago sativa） varieties， it is more difficult to identify phenotypes by traditional morphological methods. In the process of alfalfa breeding， molecular markers can greatly improve the breeding efficiency and variety identification. Reverse transcriptional transposon long terminal repeat sequence （LTR） are widely distributed in plant genomes. Molecular markers based on LTR are characterized by rich polymorphism and high information content， and are widely used in species identification and germplasm resource diversity evaluation. In this study， a large number of inter-retrotransposon amplified polymorphism （IRAP） markers were identified and designed at the genome-wide level in Medicago truncatula and used to analyze the genetic diversity of 40 alfalfa germplasm lines from within China and abroad. This approach produced 431 IRAP primers according to the design standards and the chromosomal location， from which 69 pairs of IRAP primers were obtained. A total of 325 alleles were amplified by 37 pairs of polymorphic primers； The average number of alleles was 8.8 per marker. Percentage of polymorphic bands （PPB） ranged from 50% to 100% with a mean of 79.9%. The polymorphic information content （PIC） ranged from 0.34 to 0.88， with an average of 0.69. Based on the genetic similarity coefficient （GS）， unweighted pair-group method with arithmetic means （UPGMA） was used for cluster analysis of the tested varieties. With 0.82 as the threshold， 40 samples of the tested germplasm lines could be divided into 4 categories. The grouping results were relatively consistent with geographic distribution information and STRUCTURE analysis of the 40 lines. In this study， IRAP molecular markers were developed at the whole genome level in M. truncatula for the first time and then evaluated and applied to investigate germplasm lines of alfalfa. A large number of the IRAP molecular markers obtained could provide technical support for the identification， protection and genetic background analysis of alfalfa varieties.

Key words: alfalfa, retrotransposon, IRAP marker, genetic diversity