南方农业学报2017，Vol.48Issue(7)：1304-1309,6.DOI:10.3969/j.issn.2095-1191.2017.07.29
港口航道致病性细菌检测微阵列基因芯片的研制
Development of microarray gene chip for detection of pathogenic bacteria in port and waterway
摘要
Abstract
[Objective]Gene chip to detect pathogenic bacteria in port and waterway was developed in order to lay a foundation for gene chip detection technology targeting port and waterway. [Method]The genomic DNA of the target strains were extracted by conventional methods. 16S rDNA universal primer and conserved region primers of gyrB gene were used for PCR amplification. Oligonucleotide probe designed on target fragments obtained by amplification was carried out using AlleleID 6.0 and Array Designer 4.25. Through PCR screening and validation, amino-modified target probes were spotted on aldehyde slide via microarrayer. Chip hybridization conditions were optimized, and was applied to detection of water and sediment samples from port and waterway. In this way, detection effects of microarray gene chip were validated.[Result]The optimized chip hybridization conditions were: probe concentration 10 μmol/L, ultraviolet crosslinking time 2.0 h and hybridization temperature 65 ℃. It increased sensitivity of gene chip detection, and was of high speed, high throughput and accuracy compared with traditional detection method. The microarray gene chip could detect seven pathogenic strains from port and waterway: Vibrio cholerae, Enterobacter cloacae, V. alginolyticus, V. harveyi, V. parahemolyticus, Aeromonas hydrophila and V. vulnificus. No specific hybridization was detected. [Conclusion]Micro-array gene chip detection technology targeting pathogenic bacteria in port and waterway is highly specific, sensitive, rapid and convenient. It can be used for marine environmental monitoring and marine products quality and safety detection in port and their waterway and surrounding areas.关键词
致病性细菌/港口航道/基因芯片/检测Key words
pathogenic bacteria/port and waterway/gene chip/detection分类
农业科技引用本文复制引用
郭微微,邱军强,何培民,管峰,杨先乐..港口航道致病性细菌检测微阵列基因芯片的研制[J].南方农业学报,2017,48(7):1304-1309,6.基金项目
国家海洋局海洋公益性行业科研专项项目(201105007) （201105007）
