广东海洋大学学报2025，Vol.45Issue(6)：28-34,7.DOI:10.3969/j.issn.1673-9159.2025.06.004
基于qPCR-HRM技术的诸氏鲻虾虎鱼遗传性别鉴定方法
Genetic Sex Identification in Mugilogobius chulae Based on qPCR-HRM Technology
摘要
Abstract
[Objective]This study aims to establish a reliable approach for genetic sex identification in Mugilogobius chulae when gonadal differentiation or external sex characteristics are absent,and expand the application scope of this species as a model fish in the ocean[Method]Based on two highly homologous but sex-specific transcripts(Mc15468 and Mc15469)screened from gonadal full-length transcriptome data,primers were designed flanking their variable regions of the two sequences.Using the methods of PCR amplification molecular cloning and fluorescence quantitative PCR combined with high resolution melting curve(qPCR-HRM),specific DNA molecular markers of M.chulae were identified,and a rapid method for identifying the genetic sex of M.chulae was established.This method was used to verify the genetic sex identification of 100 laboratory standardized cultured and 65 field collected M.chulae.[Result]PCR amplification and molecular cloning revealed that these primers produced two DNA fragments(101 bp and 107 bp)in males,but only one fragment(107 bp)in females,which could not be distinguished by conventional agarose gel electrophoresis.Using the method of qPCR-HRM,males exhibited two distinct melting peaks,whereas females displayed only one,enabling clear discrimination between sexes.The accuracy of the qPCR-HRM method was validated in 100 laboratory-reared and 65 wild-caught individuals,with results fully consistent with gonadal histology.[Conclusion]The established qPCR-HRM method provides a precise and rapid tool for genetic sex identification in M.chulae prior to gonad differentiation or visible sex traits,thereby facilitating studies on sex determination mechanisms and enhancing the species'utility as a marine model in environmental monitoring and ecological assessment.关键词
诸氏鲻虾虎鱼/高分辨率熔解曲线(HRM)/DNA分子标记/性别鉴定Key words
Mugilogobius chulae/High Resolution Melting curve(HRM)/DHA molecular markers/sex identification分类
生物科学引用本文复制引用
董忠典,高佳豪,李建军,曾进,郭昱嵩,王中铎,张宁..基于qPCR-HRM技术的诸氏鲻虾虎鱼遗传性别鉴定方法[J].广东海洋大学学报,2025,45(6):28-34,7.基金项目
广东省自然科学基金(2025A1515010893) （2025A1515010893）
国家自然科学基金面上项目(42477288) （42477288）
广东海洋大学水产优青B类项目 （）
广东海洋大学本科生创新创业项目(CXTD2023) （CXTD2023）
广东海洋大学大学生创新创业项目(S202410566009). （S202410566009）
