生物技术通报2024，Vol.40Issue(5)：94-102,9.DOI:10.13560/j.cnki.biotech.bull.1985.2023-0967
适于多物种的通用尾巴序列设计及通用体系的建立
Design of Universal Tailed-sequence and Establishment of the Universal System Suitable for Multiple Species
摘要
Abstract
[Objective]Fluorescent capillary electrophoresis,for its high detection throughput and resolution,was in extensive applications in various scenarios,including individual identification,variety discrimination,and species identification.The tailed-sequence offers an efficient solution for the extensive application of the fluorescent capillary electrophoresis platforms.To address the limitation of existing tailed-sequences in meeting the demands of multi-species applications,this study developed an efficient Universal Tailed-Sequence(UTS)design tool based on encoding translation technology.Utilizing this tool,UTS were designed to construct a versatile PCR system and protocol suitable for multiple crops,thereby enhancing the throughput and flexibility of fluorescent electrophoresis.[Method]An efficient UTS design tool was designed based on encoding translation technology and 3 755 commonly used Chinese characters were encoding translated with filtering criteria including GC content,hairpin,and the self-dimer annealing temperature.The BLAST tool was used to assess the homology of UTS generated by the design tool on various biological genomes.The UTS was experimentally evaluated on crops such as maize,tomatoes,peppers,and watermelons,constructing a species-universal experimental system and protocol.[Result]The design tool,through encoding translation and screening,generated a total of 7 436 833 high-quality candidate UTS,accounting for 52.74%of all character combinations.Six selected UTS were demonstrated better specificity across 20 crop genomes by BLAST results compared to M13.With optimization of the general primer amplification procedure,the success rate of general primer amplification across multiple species reached or exceeded 95%,showing strong stability.[Conclusion]This study utilized encoding translation technology to develop UTS and paired them with a species-universal amplification system and protocol,which may provide a high-throughput,cost-effective universal detection method for fluorescent electrophoresis.关键词
PCR/荧光电泳/通用引物/引物设计Key words
PCR/fluorescence electrophoresis/universal primer/primer design引用本文复制引用
孙擘,王蕊,霍永学,葛建镕,匡猛,王凤格..适于多物种的通用尾巴序列设计及通用体系的建立[J].生物技术通报,2024,40(5):94-102,9.基金项目
北京市农林科学院科技创新能力建设专项(KJCX20230301),北京市农林科学院科研创新平台建设专项(PT2023-34) （KJCX20230301）
