應用FISH-AFLP技術分析105份櫻桃種質資源的親緣關系

陳新　徐麗　宗曉娟　王甲威　譚鉞　朱東姿　魏海蓉　洪坡　劉慶忠

摘要：為建立櫻桃種質資源的FISH-AFLP技術體系并進行親緣關系分析，本研究以國內外搜集的105份櫻桃種質為對象，經DNA提取、EcoRⅠ/MseⅠ雙酶切，再進行FISH-AFLP反應。擴增結果顯示，篩選得到8對EcoRⅠ/MseⅠ多態性引物，可擴增出1 148條譜帶，平均多態性比率98.61%。聚類分析和遺傳多樣性分析結果顯示，105份櫻桃栽培品種的遺傳相似性系數為0.54～0.89，當閾值為0.68時，可分成7個AFLP群;有效等位基因數、基因多樣度、Shannon信息指數分別為1.52、0.30、0.45，具有較高的遺傳多樣性。本研究結果可為今后櫻桃品種鑒別和遺傳多樣性研究等提供理論指導。

關鍵詞：櫻桃;FISH-AFLP;遺傳多樣性;親緣關系

中圖分類號：S662.5文獻標識號：A文章編號：1001-4942（2019）11-0022-06

Genetic Relationship Analysis of 105 Accessions of

Cherry Germplasms by FISH-AFLP Technology

Chen Xin， Xu Li， Zong Xiaojuan， Wang Jiawei， Tan Yue，

Zhu Dongzi， Wei Hairong， Hong Po， Liu Qingzhong

（Shandong Institute of Pomology/Shandong Provincial Key Laboratory of

Fruit Tree Biotechnology Breeding， Taian 271000， China）

Abstract The objective of this study was to establish FISH-AFLP analysis system for cherry and analyze the genetic relationship of the main cultivars. One hundred and five cherry cultivars were selected as experiment materials. The high-quality genomic DNA was extracted and the purified DNA samples were digested with EcoRⅠ/MseⅠ;after ligation reaction， the samples were used to perform FISH-AFLP detection. The amplification results showed that eight pairs of EcoRⅠ/MseⅠ polymorphic primers were screened and 1 148 bands were obtained. The average percentage of polymorphic bands was 98.61%. The results of cluster analysis and genetic diversity analysis showed that the genetic similarity coefficient of the 105 cherry cultivars varied from 0.54 to 0.89. All the samples were divided into 7 groups when the threshold value was 0.68. The numbers of effective alleles， gene diversity and Shannon information index of the main cultivars were 1.52， 0.30 and 0.45， respectively， indicating high genetic diversity. This study could provide theoretical guidance for the identification and genetic diversity research of cherry varieties in the future.

Keywords Cherry; FISH-AFLP; Genetic diversity; Genetic relationship

櫻桃為多年生木本果樹，屬薔薇科櫻桃屬。目前世界上作為果樹栽培的櫻桃有4個種，品種繁多，僅栽培品種就超過2 000個。隨著我國新品種的不斷選育以及國外品種的引進，基于表型性狀標記鑒定不同品種以及確定品種分類地位及相互之間的親緣關系越來越困難。DNA分子標記在櫻桃上的應用已有報道。其中，RAPD[1-3]、SSR[4，5]皆曾用于櫻桃種質資源的研究，但揭示的多態性位點較少;國外研究者曾利用cpSCAR分析櫻桃品種間的遺傳關系和多樣性[6]，但多集中在親緣關系較近的歐洲甜櫻桃品種上，尤其是一些老品種。

AFLP（amplified fragment length polymorphism，擴增片段長度多態性）檢測DNA多態性的方法，于1995年以論文形式發表[7]，是通過限制性內切酶片段的不同長度檢測DNA多態性的一種DNA分子標記技術。AFLP具有多態性強、譜帶豐富和靈敏、分辨率高、快速高效、重復性好等特點，是一種較為理想的分子標記技術[8]，可用于檢測種及種以下水平上的差異。……