NEXT GENERATION SEQUENCING METHOD OF ZOOBENTHOS CYTOCHROME OXIDASE SUBUNIT I GENE AND USE THEREOF

• Main Authors: YIXUAN REN, SHENGJIE CAO, JIANING LIN, SHUPING WANG, XIAOBO JIA, HUI WANG, SEN DING, YUAN ZHANG, RUI XIA, WEI SUN
• Format: Patent
• Language: English
• Published: 19.07.2022
• Subjects: BEER; BIOCHEMISTRY; CHEMISTRY; COMPOSITIONS OR TEST PAPERS THEREFOR; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL ORENZYMOLOGICAL PROCESSES; ENZYMOLOGY; MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEICACIDS OR MICROORGANISMS; METALLURGY; MICROBIOLOGY; MUTATION OR GENETIC ENGINEERING; PROCESSES OF PREPARING SUCH COMPOSITIONS; SPIRITS; VINEGAR; WINE

The present invention relates to the technical field of zoobenthos identification, and in particular to a next generation sequencing (NGS) method of a zoobenthos cytochrome oxidase subunit T (COT) gene and use thereof. The method 5 includes the following steps: Sl, amplifying the COT gene by polymerase chain reaction (PCR) with a DNA sample of a mixed zoobenthos sample as a template; SZ, amplifying a PCR.product obtained in step Sl as a template by PCR, to obtain a PCR product as a DNA library, where an amplified primer pair includes a 10 pyroseguencing-NGS adapter and each sample corresponds to one pyroseguencing-NGS adapter; and S3, conducting guality detectioncm1aresultinglflwxlibraryy andselectingeagualified DNA_library to conduct emPCR.amplification.and.NGS. The method obtains a high-throughput full-length (~658 bp) COT gene 15 sequence with read length covering large benthic communities by all-at-once sequencing, providing a large taxonomic resolutionandavoidingfalsepositivesdueixnexcessivelyhigh throughput.