Construction of miR-142-5p overexpression lentiviral vector and its impact on mouse intestinal epithelial cells
10.16303/j.cnki.1005-4545.2025.04.08
- VernacularTitle:miR-142-5p过表达慢病毒载体的构建及对小鼠肠道上皮细胞的影响
- Author:
Yicheng SUN
1
;
Jing WANG
1
;
Yushu HE
1
;
Zijian DONG
1
;
Jing DING
1
Author Information
1. 吉林大学 动物医学学院人畜共患传染病重症诊治全国重点实验室/人兽共患病研究所/人兽共患病研究教育部重点实验室,吉林长春 130062
- Publication Type:Journal Article
- Keywords:
miR-142-5p;
lentivirus;
intestinal epithelial cells
- From:
Chinese Journal of Veterinary Science
2025;45(4):678-684
- CountryChina
- Language:Chinese
-
Abstract:
To investigate the impact of miR-142-5p on intestinal epithelial cells,an expression vector for overexpressing miR-142-5p lentivirus was constructed and administered to mice to observe pathological changes in their colon tissue.The miR-142-5p gene was integrated into the green fluo-rescent lentiviral vector plenti-CMV/TO eGFP-Puro(plenti),and the resulting recombinant vec-tor was named plenti miR-142-5p following confirmation through sequencing.293T cells were transfected with the recombinant plasmid and packaging-assisted plasmid,yielding a packaged miR-142-5p overexpressing lentiviral vector(LV-miR-142-5p).The recombinant lentivirus suspen-sion was collected and concentrated via ultrafast centrifugal precipitation.Virus titer was deter-mined using real-time PCR.The virus suspension was then injected into the tail vein of 6-8-week-old BALB/c mice,after which their colonic tissues were dissected for HE staining and microscopic observation.The results showed that miR-142-5p homologous recombination into plenti vector and sequencing results were consistent with the expected sequence.plenti-miR-142-5p was co-transfect-ed with the packaging helper plasmid in 293T cells,and the successful transfection was confirmed based on the green fluorescence expression,and the real-time PCR results showed that the lentiviral titer was 1.23 × 109 IU/mL.Colon tissue slices from infected mice exhibited compromised colon integrity and significant inflammatory response.It was proved that LV-miR-142-5p was suc-cessfully constructed,and miR-142-5p caused damage to intestinal epithelial cells of mice.
