桃PpCBF2基因克隆及其响应低温表达特性研究

Cloning of PpCBF2 gene in peach and its response to low temperature expression

为探讨桃品种间抗寒差异的分子基础,以野生毛桃(Prunus persica)和17个桃品种和品系为试验材料,通过PCR与RT-PCR克隆PpCBF2基因;并利用qRT-PCR分析‘早红霞’和‘大久保’叶片中PpCBF2在低温下的表达特性。结果表明:1)毛桃PpCBF2基因的DNA和cDNA全长序列均为894bp,开放阅读框为690bp,编码229个氨基酸并且含有CBF家族的AP2/EREBP功能域;2)供试的17个桃品种和品系的PpCBF2基因核苷酸序列全长和编码氨基酸数目与毛桃相同,但有9个品种发生1~2个氨基酸位点突变:其中‘白芒蟠桃’、‘81715’和‘大久保’的突变发生在N-豆蔻酰化位点或酪蛋白激酶II磷酸化位点;‘Italia 3’、‘寒露’和‘B6832’的突变发生在AP2/EREBP功能域;3)‘早红霞’和‘大久保’PpCBF2基因均被低温诱导,品种间表达水平有显著差异。综上,该基因在桃品种间进化较为保守,但少数品种基因功能位点发生了突变;该基因被低温显著诱导,与抗冷性有关。

In order to explore the molecular basis of the differences in cold resistance among peach varieties,wild peach(Prunus persica)and 17 peach varieties and lines were used as test materials.The PpCBF2 gene was cloned by PCR and RT-PCR.The expression characteristics of PpCBF2 at low temperature in‘Zaohongxia’and‘Okubo’leaves were analyzed by qRT-PCR.The results showed that:1)the full-length DNA and cDNA sequences of PpCBF2 gene were 894 bp,the open reading frame was 690 bp,encoding the 229 amino acids and containing the AP2/EREBP domain of the CBF family;2)The full-length nucleotide sequence and the number of encoded amino acids of the PpCBF2 gene of the strain were the same as those of the Prunus persica,but there were 1-2 amino acid site mutations in 9 varieties:The mutations of‘Baimangpantao’,‘81715’,and‘Okubo’occurred at N-myristoylation site or the phosphorylation site of casein kinase II;mutations in‘Italia3’,‘Hanlu’,and‘B6832’occurred in the AP2/EREBP domain;3)PpCBF2 of‘Zaohongxia’and‘Okubo’were induced by low temperature,and there were significant differences in expression levels.In conclusion,PpCBF2 gene was relatively conservative among peach varieties,but a few varieties had mutations in gene functional sites;this gene was significantly induced by low temperature and was related to cold resistance.