The Comparing Analysis Of The Genomic Difference Of Grape Eearly Bud Mutation And Its Parents

Bud mutation is a common phenomenon in grape. ’Kyoho’ and ’Jingya’ are currently two mainly cultivated varieties. ’Fengzao’ and ’Luopuzaosheng’ are their corresponding bud mutant. Compared with the parent, the maturity date of ’Fengzao’ and ’Luopuzaosheng’ are earlier about 30 d and 20 d than their parents.The purpose of this study was to compare the genomic differences between the two precocious mutant varieties and their parents on the DNA level from L Ⅱand L Ⅰ+L Ⅱlayer by i PBS and MSAP markers. The main results are as follows:1. iPBS mark was employed to detect the difference based on the retrotransposon characteristics. Three out of fifteen iPBS primers showed no different bands between ’Fengzao’ and the ’Kyoho’. The remaining twelve have shown distinctive bands. Nine primers showed that the LⅡ layer of ’Fengzao’ mutated, LⅠ + LⅡ layer also mutated; three primers showed that the LⅡ layer of ’Fengzao’ mutated, LⅠ + LⅡ layer did not mutate. Six out of fifteen iPBS primers showed no different bands between ’Luopuzaosheng’ and ’Jingya’, the remaining nine have shown different bands. Six primers showed that the LⅡ layer of ’Luopuzaosheng’ mutated, LⅠ + LⅡ layer also mutated; three primers showed that the LⅡ layer of ’Fengzao’ mutated, LⅠ + LⅡ layer did not mutate. In all, the LⅡ layer of precocious bud mutation mutated, but the LⅠ layer can not be determined whether mutated or not.2. The difference analysis results of parentand the mutant by MSAP are as follows:(1) The methylation difference is mainly concentrated in type Ⅱ(31 <70, 55 <91) between mutant and parent; that is, the main cause of differential methylation is CCGG sites of cytosine demethylation occurred between mutant and parent. In addition, the methylation rates of the mutant were generally lower than the parental methylation rate(20.5% <26.3%, 20.1% <28.1%), it further illustrate that the mutant is occured due to the sites of CCGG cytosine demethylation, resulting in precocious formation.(2) The methylation variance level analysis inleaf, peel, root and flesh of mutant and parental, discovered the different degrees of methylation at the site; ’Fengzao’ demethylation occurred mainly in the roots, while ’Luopuzaosheng’ mainly in the pulp and root.(3) The methylation variance level analysis of different organs of mutant and parent;revealed that the CCGG sites of methylation patterns of the leaves and roots have changed in ’Fengzao’ and ’Luopuzaosheng’, showing external methylation reduced, internal methylation increased; the demethylation sites of CCGG in the peel of ’Fengzao’ turned out demethylation, while methylation in ’Luopuzaosheng’ peel did not changed significantly; the methylation patterns of the flesh of ’Fengzao’ and ’Luopuzaosheng’ occured different degrees of change. The different degrees of methylation in different organs of bud mutation and parents, indicated that DNA methylation may be involved in the differentially expressed organization in the process of the mutant.(4) The difference analysis of LⅡ layer in the mutant parental, by comparing the difference of bud mutation and parental methylation rates,outside methylation rates, methylation rates, suggested that the methylation rate of bud mutant is significantly less than that of parent, indicating that cytogenetical demethylation occurred in the LⅡ layer CCGG sites of bud mutant; the outside methylation rate of bud mutant is significantly lower than the parent indicating that the LⅡ layer CCGG external site of bud mutant occurred demethylation, that is, the LⅡlayer of bud mutant mutated; inter methylation rate of ’Fengzao’ is significantly higher than that of ’Kyoho’, showing that methylation occurred in the LⅡ layer CCGG inter site of ’Fengzao’; while ’Luopuzaosheng’ inter methylation rate is bigger than ’Jingya, showing that the LⅡ layer CCGG inter site of ’Luopuzaosheng’ occurred methylation,but the difference is not obvious.(5) The difference analysis of LⅠ+L Ⅱlayer by comparing the difference of bud mutation and parental methylation rates,outside methylation rate, methylation rates suggested that the methylation rate of bud mutant is less than parental methylation rate, indicating that the LⅠ+L Ⅱlayer of bud mutant is demethylation, but not obvious; outer methylation rate of ’Fengzao’ is significantly less than ’Kyoho’ o, showing that demethylation occurred in the LⅠ+L Ⅱlayer CCGG outer site of ’Fengzao’; while ’Luopuzaosheng’ alter methylation rate is bigger than ’Jingya’, showing that methylation occurred in the LⅠ+L Ⅱlayer CCGG outer site of ’Luopuzaosheng’; the inter methylation difference of ’Fengzao’ and ’Kyoho’ is not obvious; while ’Luopuzaosheng’ inter methylation rate is less than that of ’Jingya’, showing that demethylation occurred in the LⅠ+L Ⅱlayer CCGG inter site of ’Luopuzaosheng’,but the difference is not obvious.The difference analysis of bud mutant and parent by i PBS and MSAP techniques showed that the bud mutation changed due to the mutation of the L Ⅱlayer whether LⅠlayer changed or not, that is unclear.