文献类型：期刊文献

中文题名：基于流式细胞仪鉴定树莓倍性方法的建立及应用

英文题名：Establishment and Application of a Method for Identifying Raspberry Ploidy Based on Flow Cytometry

作者：青梦瑶[1] 田林[2] 李迎超[3] 史瑞基[1] 张瑞杰[1] 郑奕宸[1] 孙权[1] 李寒[4] 顾玉红[1]

第一作者：青梦瑶

机构：[1]河北农业大学生命科学学院,保定071001;[2]中国林业科学研究院林木遗传育种国家重点实验室,北京100091;[3]中国林业科学研究院林业研究所,北京100091;[4]河北农业大学林学院,保定071001

年份：2025

卷号：41

期号：11

起止页码：228-235

中文期刊名：生物技术通报

外文期刊名：Biotechnology Bulletin

收录：;北大核心:【北大核心2023】；

基金：河北省重点研发计划项目(20326338D)。

语种：中文

中文关键词：树莓;细胞;倍性检测;流式细胞术;叶片;根;愈伤组织;快速鉴定

外文关键词：raspberry;cell;ploidy identification;flow cytometry;leaves;roots;callus tissue;rapid identification

分类号：S663.2

摘要：【目的】建立一套高效易行检测树莓细胞倍性的方法,鉴定不同树莓种质的细胞倍性,为树莓倍性育种和杂交育种奠定理论基础。【方法】以树莓叶片为材料,对不同的解离液、叶片来源、叶龄、叶片的保存和运输方式等条件进行优化,建立流式细胞术检测树莓细胞倍性的方法,并使用此方法检测黑树莓、红树莓、黄树莓等不同树莓种质叶片、根以及愈伤组织的细胞倍性。【结果】流式细胞仪法检测树莓叶片细胞倍性时,解离液Ⅵ(WPB)制备的细胞核悬浮液上机检测效果最好,收集到的细胞核数最多,峰图最好;田间苗和组培苗嫩叶制备的细胞核悬浮液检测效果最好,成龄叶次之;把叶片夹在2层润湿滤纸中间并放于塑料平皿后,置于低温泡沫箱中,直接带着样品上机检测效果更好,将叶片从植株剪下后直接浸泡至装有无菌水的50 mL离心管中,再邮寄样品上机检测的效果次之;流式细胞仪检测叶片的直方图主峰位于荧光强度值10000附近,用此检测方法鉴定了15个树莓种质叶片细胞的倍性均为二倍体,鉴定了‘龙园双丰’‘波尔卡’的根和愈伤组织的细胞倍性也均为二倍体。【结论】建立了一套能够检测不同发育时期和不同生长环境下树莓叶片和根以及愈伤组织细胞倍性的方法,并用此方法检测了黑树莓、红树莓和黄树莓共15个不同种质树莓倍性均为二倍体。
【Objective】To establish a set of efficient and easy-to-operate methods for detecting the ploidy of raspberry cells,and then use this method to identify the ploidy of different raspberry germplasms,laying a theoretical foundation for ploidy breeding and hybrid breeding of raspberry.【Method】Taking raspberry leaves as the material,different dissociating solutions,leaf sources,leaf ages,leaf preservation and transportation methods and other conditions were optimized to establish a method for detecting the ploidy of raspberry cells by flow cytometry.This method was used to detect the cell ploidy of leaves,roots and callus of different raspberry germplasms including black raspberry,red raspberry and yellow raspberry.【Result】The nuclear suspension prepared with dissociating solution VI(WPB)had the best detection effect when using flow cytometry to detect the ploidy of raspberry leaf cells,with the most collected nucleus and the best peak graph.The nuclear suspension prepared from the young leaves of field-seedlings and tissue-culturing seedlings had the best detection effect,followed by that from adult leaves.The best detection effect was achieved by placing the leaves between two layers of moist filter paper and then putting them in a plastic petri dish,and then placing the plastic petri dish in a low-temperature foam box for direct detection with the sample.The second-best effect was achieved by cutting the leaves from the plant and directly soaking them in a 50 mL sterile water-filled centrifuge tube before mailing the sample for detection.The main peak of the flow cytometry histogram of the leaves was located near the fluorescence intensity value of 10000.Using this detection method,the ploidy of 15 raspberry germplasm leaf cells was all determined to be diploid,and the ploidy of the roots and callus of‘Longyuan shuangfeng’and‘Polka’was also determined to be diploid.【Conclusion】This study established a method capable of detecting the cell ploidy of raspberry leaves,roots,and calli at different developmental stages and under different growth environments.Using this method,a total of 15 raspberry germplasms,including black raspberries,red raspberries,and yellow raspberries,were tested,and all were found to be diploid.