苹果梨变异单系生物学特性调查比较及RAPD多态性分析

【作者】 刘迪；

【导师】 曲柏宏；

【作者基本信息】 延边大学 ， 果树学， 2007， 硕士

【摘要】 本试验以苹果梨及其11个变异单系为材料（分别采自延边州农业科学研究院、珲春市农垦特产局和延边大学农学院），通过田间调查比较，果实品质分析，初步筛选出与苹果梨存在一定差异的优良单系。并以苹果梨及其变异单系的成熟嫩叶为试材提取DNA，对其进行RAPD多态性分析。结果表明：苹果梨在大面积栽培过程中发生了遗传变异，供试苹果梨单系在果实品质方面表现出一定的差异，部分变异单系个别性状或综合性状优于苹果梨。通过分析，综合性状表现较好的变异单系是P₁、P₅、P₇、P₈。采用CTAB改良法快速提取到了较高质量苹果梨及其变异单系基因组总DNA用于RAPD扩增，并在RAPD反应中筛选出18个10碱基随机引物，它们的多态性百分率大多在67.7以上，确定了RAPD-PCR苹果梨及其变异单系最适反应体系为：模板DNA浓度为15ng／μL，Mg²⁺浓度为1.0mmol／L，Taq DNA聚合酶1U，dNTP的浓度为1.0mmol／L，10碱基引物浓度为15pmol，10×缓冲液浓度为2.5mmol／L，其余用重蒸馏水补充。扩增程序为：94℃预变性5min，94℃变性1min，37℃退火1min，72℃延伸1.5min，共45个循环：最后72℃延伸7min；终止温度为4℃。建立了苹果梨及其11个变异单系的指纹图谱。找到了P₂和P₇两个变异单系的特征标记，其中P₇为大果型变异；苹果梨11个变异单系可以用一个引物的一条带和苹果梨区分开。更多还原

【Abstract】 Applying Pingguoli（Pyrus ussuriensis var.ovoidea_XP.Pyrifolia）and its eleven varied strains as testing materials,several good strains of synthetical properties were screened out of the others through the investigation of biological characteristics and the analysis of fruits quality. On base of these, using the ripe and fresh leaves of Pingguoli and its eleven variation strains, genome DNA was abstracted and polymorphic of RAPD was studied. The results showed that:Some genetic variation occurred in the course of spreeding and cultivating of Pingguoli .In all varied strains, there were some divergences in quality of fruit, and the exceptional or synthetical characters of some varied strains were better than Pingguoli .By identifying and analyzing, four varied strains whose synthetical character were better were P₁、P₅、P₇、P₈.An improved CTAB method could extracted high-quality DNA from gene group of Pingguoli and its variation strains and filter eighteen 10 random primer in RAPD amplification procedures, their polymorphism rate were over 67.7. An optimal reaction systern for RAPD-PCR was: template DNA 15ng/u L, Mg²⁺ 1.0mmol/L, Taq DNA polymerase 1 unit, dNTP 1.0mmol/L, 10-code primer 15pmol, 10×buffer 2.5mmol/L, others complement with ddH₂O. The amplification procedures was: initial denaturation at 94^C for 5 minutes; denaturation at 94^C for 1 minute, annealing at 37 for 1 minute, elongation at 72^C for 1.5minutes, 45 cycles; final elongation at 72^C for 7 minutes, stop at 4^C.Fingerprints of Pingguoli and its eleven varied strains were established .The marker of P₂和P₇ were founded, 11 variation strains can be distinguished from from Pingguoli in one primer.更多还原