Establishment method of cLYZ/hTLF double-antibacterial gene recombinant adenovirus, and recombinant adenovirus and application thereof

• Main Authors: GONG JIANFANG, SHANG CUILING, JIN TIANMING, GAO KEKE, MENG JIALI, LI XIAOHUI, LIN JING, LI XIN, ZHANG DONGCHAO
• Format: Patent
• Language: Chinese; English
• Published: 12.10.2016
• Subjects: BEER; BIOCHEMISTRY; CHEMISTRY; COMPOSITIONS THEREOF; CULTURE MEDIA; ENZYMOLOGY; HUMAN NECESSITIES; HYGIENE; MEDICAL OR VETERINARY SCIENCE; METALLURGY; MICROBIOLOGY; MICROORGANISMS OR ENZYMES; MUTATION OR GENETIC ENGINEERING; PREPARATIONS FOR MEDICAL, DENTAL, OR TOILET PURPOSES; PROCESSES USING MICROORGANISMS; PROPAGATING, PRESERVING OR MAINTAINING MICROORGANISMS; SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS ORMEDICINAL PREPARATIONS; SPIRITS; VINEGAR; WINE

The invention discloses an establishment method of a cLYZ/hTLF double-antibacterial gene recombinant adenovirus, and a recombinant adenovirus and application thereof. The method comprises the following steps: respectively amplifying cLYZ gene and hTLF gene from grouper head-kidney tissues and human milk somatic cell total RNAs, cloning the obtained goal genes into a pDC315 shuttle vector, co-transfecting HEK293 cells through liposome by using the target-gene-containing shuttle plasmid and adenovirus framework plasmid, and packaging to obtain the high-titer recombinant adenovirus; and evaluating the combined inhibiting actions of the cLYZ gene and hTLF gene on the milk cow mastitis primary pathogens under the adenovirus mediated conditions. 本发明公开了种cLYZ和hLTF双抑菌基因重组腺病毒的构建方法及重组腺病毒和应用，从石斑鱼头肾组织和人乳体细胞总RNA中分别扩增出cLYZ基因和hLTF基因，将获得的目的基因克隆到pDC315穿梭载体中，将含有目的基因的穿梭质粒与腺病毒骨架质粒经脂质体共转染HEK293细胞，包装获得高滴度的重组腺病毒；评价cLYZ基因和hLTF基因在腺病毒介导下对奶牛乳房炎主要致病菌的联合抑制作用。