“云新高原”核桃及其亲本的叶绿体DNA差异分析

Comparison of Chloroplast DNA among Artificial Hybrid Walnut 'Yunxingaoyuan' and Its Parents

【目的】研究核桃人工种间杂交种云新高原与其亲本(母本为漾濞泡核桃,父本为云林A7)的叶绿体DNA的差异,了解核桃叶绿体DNA的遗传规律,并挖掘多态性高的叶绿体基因片段。【方法】基于NCBI数据库中深纹核桃和核桃叶绿体全基因组序列的比对和分析,针对其中的变异位点设计引物,进而对云新高原及其母本漾濞泡核桃、父本早实核桃云林A7的目的基因片段进行多态性检测。【结果】深纹核桃与核桃的叶绿体基因组存在14个变异位点、30个碱基的差异,其中7个为核苷酸多态性变异(SNP),7个为碱基插入/缺失变异(InDel)。对云新高原及其亲本的这14个位点及其附近片段进行检测,发现三者有33个变异位点,云新高原与其母本在其中21个位点上保持相同碱基,占全部变异位点数的64%;云新高原与其父本在余下12个位点上保持相同碱基,占全部变异位点数的36%。这33个变异位点分属11个基因或基因间隔区片段,可以分为11个单倍型,云新高原的这11个单倍型中有10个与母本相同,1个为双亲的嵌合体。【结论】核桃叶绿体DNA的遗传方式为以母系遗传为主的双亲遗传;核桃ndhF-rpl32DNA片段存在多个变异位点,是多态性高的叶绿体基因片段,可广泛应用于核桃系统发育、群体遗传和种内品种鉴定等研究。

【Objective】The differences of chloroplast DNA among artificial interspecific hybrid walnut,Juglans sigillata × J.regia cv.＇Yunxingaoyuan＇and its parents（its female parent is Juglans sigillata cv.＇Yangpao＇ and its paternal parent is J.regia cv.＇Yunlin A7＇） were studied in this paper,to understand the genetic rule of walnut chloroplast DNA and to mine highly variable chloroplast DNA regions.【Method】The downloaded chloroplast genome sequences of J.sigillata and J.regia from NCBI database were aligned and analyzed.The primers were designed based on the different variation positions.Then the labeling regions among＇Yunxingaoyuan＇and its parents were sequenced and verified for the polymorphism.【Result】The results showed that there were seven SNPs and seven InDels in choroplastic genome sequences of two Juglans species,of which total size involved 30 bases.Fourteen primers were designed,and all of them were successfully amplified and sequenced among＇Yunxingaoyuan＇and its parents.The resulting sequences demonstrated totally 33 variable sites,of which 21（64 %） sites were the same between ＇Yunxingaoyuan＇and its female parent,and 12（36 %） sites were the same between＇Yunxingaoyuan＇and its paternal parent.The 33 variable sites belonged to 11 genes or intergenic segments,and could be divided into 11 haplotypes,of which 10 haplotypes were identical to the female parent,and remaining 1 had the haplotypes of both parents.【Conclusion】According to the plastid inheritance law,the analysis evidences indicated that chloroplast inheritance in Juglans is maternal dominantly and biparental subordinately pattern.The ndhF-rpl32 fragment exist multiple variation sites,so it is excellent candidate for future phylogenetics,population genetics and variety identification studies.