关键词: 越桔属, 蓝莓, 栽培品种, DNA条形码, 遗传分化

Abstract:

Objectve: The purpose of this study was screening the suitable DNA barcodes for identifying cultivars, which were used for studying the discrimination and the heredity differentiation of blueberry (Vaccinium) cultivars. Method: Eight DNA barcodes (rpoB, psbA-trnH, ycf5, rbcL, rpoC, matK, ITS and ITS2) of 108 individuals from 40 blueberry cultivars were amplified and sequenced. The amplification efficiency, sequencing success rate, genetic distance and barcoding gap were analyzed, and the calculations of genetic distance were verified using the Wilcoxon test. Finally, cluster analysis of blueberry cultivars was performed by N-J tree method. Result: Among the eight DNA barcodes, ITS and ycf5 could not be amplified in all samples. The amplification and sequencing rate of all the five barcodes (rpoB, psbA-trnH, rbcL, rpoC and ITS2) were 100%, except matK was 96.3% and 99.04% respectively. The rpoB sequences of all blueberry cultivars were completely consistent, so this fragment is highly conserved in blueberry. The mutation points of six barcodes were as follows:ITS2(11) > matK(4) > rbcL(3) > psbA-trnH(2) > rpoC(1) > rpoB(0). The genetic distances among or within blueberry cultivars were smaller, range from 0.000 16-0.002 58. The genetic distances were greater among cultivars than within cultivars. Barcoding gap analysis showed that, there was no obvious interval area formed in all barcodes, but according to distribution, barcodes ITS2, psbA-trnH and matK tended to be distributed at both ends, especially ITS2. Wilcoxon test showed that, the interspecific variation of barcodes ITS2 and psbA-trnH were more obvious than others, and greater intraspecific variation was found in psbA-trnH. The cluster analysis indicated that, blueberry cultivars could be divided into two groups by psbA-trnH or rpoC, three groups by rbcL or matK, and four groups by ITS2. Furthermore, the identification rate of blueberry cultivars could be improved by the combination of barcodes and with the highest success rate of 20% by ITS2+matK+rpoC+rbcL. Conclusion: The identification of blueberry cultivars by ITS2 was superior to other barcodes. A total of 40 blueberry cultivars were divided into 14 groups by the barcode combination of ITS2+matK+rpoC+rbcL, which can distinguish the major blueberry cultivars in Shandong province, such as 'Berkeley', 'Sunshineblue' and 'Northland' respectively. The barcode combination is suitable for the identification of blueberry cultivars.

Key words: Vaccinium, blueberry, cultivar, DNA barcode, genetic divergence