Identification of true hybrids of open-pollination seedling progenies in mango and genetic characterization of the progenies using SSR markers

Abstract:【Objective】As a perennial woody fruit tree, mango (Mangifera indica L.) has the problems of high genomic heterozygosity, unclear self-compatibility and hybridization affinity of most breeding parents, and low seed setting rate of artificial pollination, which brings some difficulties to the selection of parents for mango hybridization. SSR markers are mainly used in genetic diversity analysis, genetic relationship analysis and fingerprint construction in mango, but the use of SSR markers for identifying the true hybrids of mango F1 progeny has not been reported. In this study, 13 varieties were selected as parental materials, and SSR technology was used to detect the true hybrids and analyze the genetic characteristics of 1001 F1 progeny plants in order to provide theoretical guidance for clarifying the hybridization affinity of mango breeding parents and the selection of breeding parents.【Methods】The 13mango varieties were mixed and planted for natural pollination. The mature fruits were harvested from the fruiting mother tree. The mother varieties included 12 varieties such as Guifei, Tainong No. 1, Jinhuang, Juile, Nan Doc Mai, Yexiang, Villard, R2E2, Tangmi, Hongyu, A61 and Keitt. The seedlings were obtained after sowing. 100 mg of fresh tissue of the leaves of each seedling were taken with a puncher. The DNA was extracted by magnetic bead genomic DNA extraction kit, and the concentration and purity of DNA were detected by ultraviolet spectrophotometer and 1% agarose gel electrophoresis, the PCR products were detected by agarose gel electrophoresis (1% concentration) and capillary fluorescence electrophoresis. 200 pairs of primers were selected from the previous transcriptome data of mango for screening, and 200 pairs of primers were synthesized using each parent as the template of DNA. The screened primers were used for SSR analysis. The genetic parameters such as number of alleles (Na), number of effective genes (Ne), observed heterozygosity (Ho), expected heterozygosity (He), fixation index (F) and Shannon’s information index (I) were calculated by the Cervus software; The polymorphism information index (PIC) was calculated by the GenALEx6.0 software, and the genetic differentiation coefficient was obtained; The Structure was used to analyze the genetic structure of the population and the genetic diversity of mango F1 generation; The unweighted pair-group method with arithmetic means (UPGMA) was used to cluster the populations of mango offspring by using the selected SSR core primers.【Results】Thirteen pairs of primers with stable amplification and clear bands were screened, and 13 loci with good polymorphism and stability were determined. The PIC value was between 0.314 and 0.741, and the polymorphism of the loci was high. The number of alleles ranged from 3 to 9, with an average of 6.154. The average effective number of alleles (Ne) was 2.557, the Shannon’s information index (I) was 1.078, the observed heterozygosity (Ho) was 0.571, the expected heterozygosity (He) was 0.579, and the polymorphism information content (PIC) was 0.526; The UPGMA cluster analysis showed that when the genetic similarity coefficient was 0.50, it could be divided into three groups, which was basically consistent with the results of population structure analysis. Compared with other varieties, the alleles of mango varieties Guifei, Yexiang, and Juile were more evenly distributed in the population, the average Shannon’s information index (I) was higher, and Juile was the lowest, indicating that the genetic diversity in the Guifei population was higher and the population differentiation was the highest; The genetic differentiation between Guifei and Juile was obvious, and the genetic differentiation between the other varieties was small; The true hybrid rates of the open-pollination progenies generated from the mother plants of Guifei, Jinhuang, Keitt, Tainong No. 1, A61, Juile and Keitt were over 50%, and the true hybrid rate of Jinhuang, Tainong No. 1 and A61 was as high as 80%; In terms of expected heterozygosity and observed heterozygosity, the heterozygosity of Tainong No. 1, Keitt, Juile, Jinhuang, Hongyu and Guifei was also relatively high, and the true heterozygosity of Nan Doc Mai, R2E2, Hongyu, Villard and Yexiang, was low, which was 25.00%, 13.71%, 25.15%, 33.33% and 36.36%, respectively.【Conclusion】The genetic differentiation between Hongyu and R2E2, Hongyu and Yexiang, Hongyu and Nan Doc Mai, Juile and R2E2, Juile and Nan Doc Mai, Guifei and Nan Doc Mai, Guifei and Juile was obvious, and the genetic differentiation between the remaining varieties was small; The SSR identification of mango open-pollination generation showed that the crossing compatibility of mango varieties Tainong No. 1, Keitt, Juile, Jinhuang, Hongyu and Guifei was strong, while the crossing compatibility of Nan Doc Mai, R2E2, Hongyu, Villard and Yexiang, varieties was weak, which was easy to self-fertilize, and there were many alleles among the varieties, and the genetic diversity was rich.