Genetic relationship analysis of 19 accessions of yellow peach germplasms based on SCoT markers

Abstract:【Objective】Many yellow peach germplasms are difficult to distinguish because of their simi-lar biological morphology. SCoT (Start Codon Targeted polymorphism) molecular markers were screened to analyze the genetic diversity and genetic relationship of yellow peach germplasms to pro-vide theoretical basis for breeding new yellow peach variety and avoid the reuse of core parents and nar-row genetic background of new variety.【Methods】19 accessions of yellow peach including 18 peach cultivars and 1 excellent strain were collected, and most of them were mainstream varieties in peach in-dustry. The genomic DNA of 19 peach materials was extracted from young leaves by CTAB method.The SCoT technique was based on the single primer amplified region principle since it used a single primer as a forward and reverse primer. The SCoT primers were designed with fixed principles. Firstly,the ATG codon (+1, +2, and +3) and the base G, A, C and C, at positions +4, +7, +8, and +9 were fixed.Secondly, primers were sequences of 18 bases and ranged in GC content between 50% and 72%. 15 SCoT primers, including 10 primers beginning with name“SCoT”designed by previous researches and 5 primers beginning with the name“HSCoT”designed by this study, were selected to analyze the 19 yellow peach varieties (lines). The SCoT-PCR reaction system was 20.0 μL, and its reaction mixtures contained 1 μL 2.5 mmol·L^-1 Mg₂+, 0.4 μL 0.3 mmol·L^-1 dNTPs, 1 μL 30 mg·L^-1 DNA template, 0.5 μL10.0 μmol · L^-1 primer, 0.5 μL 0.4 U Taq DNA polymerase and 16.6 μL ddH2O. A PCR cycle reaction program was applied: an initial denaturation step at 94 ℃ for 5 min, followed by 35 cycles of 94 ℃ for 30 seconds, 48 ℃ for 30 seconds, 72 ℃ for 2 min and the final extension at 72 ℃ was held for 10 min. The PCR amplification products were separated by 1.8% agarose gels and photographed on the SAGE CREATION ChamlGel 6000 imaging system under UV light. To form a“1/0”matrix, detected PCR-amplified fragment was recorded“1”and no amplified fragment was recorded“0”at the same level of gel. The“1/0”matrix was utilized to carry out cluster analysis, genetic similarity coefficient and princi- pal coordinate analysis with Software NTSYS 2.10.【Results】15 SCoT primers were used to amplify 19 cultivars (lines) of yellow peach by PCR program. A total of 283 clear bands were amplified by 15 prim-ers, with an average of 18.86 bands amplified by each primer. Among the amplified bands, 277 bands were polymorphic, with a ratio of 97.88%. Using the Software NTSYS 2.10, the genetic similarity coef-ficients of 19 peach cultivars were caculated, they ranged from 0.186 to 0.635, with an average of 0.324. The highest genetic similarity coefficient between cultivars Jinhong and Jinchun was 0.635, and the lowest coefficient between cultivars Guantao 5 and line 1 was 0.188. The cluster analysis of 19 peach cultivars (lines) was carried out by UPGMA method, and when the genetic similarity coefficient was 0.27, the 19 cultivars could be divided into 5 groups. The first group mainly contained foreign culti-vars Golden boy 6 and Xiweihuangjin. The second group mainly contained a series of Huangjinmi culti- vars bred by Zhengzhou Fruit Research Institute. The third group mainly contained a series of Jin culti- vars bred by Shanghai Forest and Fruit Tree Research Institute. The fourth group contained two culti-vars of Jin series and one cultivar of Huangjinmi series, showing distant differences from other Jin se-ries and Huangjinmi series in the genetic background. The last group contained only one cultivar Guan-tao 5, which mainly was used for canned food processing. The principal coordinate analysis divided 19 peach cultivars (lines) into 6 groups. Compared with the cluster analysis, the extra group in the princi-pal coordinate analysis came from a division of the third group in the cluster analysis, and the other four groups were consistent.【Conclusion】19 accessions of yellow peach were divided into 5 or 6 groups by SCoT marker analysis.