期刊
POLISH JOURNAL OF VETERINARY SCIENCES
卷 19, 期 3, 页码 619-625出版社
POLSKA AKAD NAUK, POLISH ACAD SCIENCES, UNIV WARMIA & MAZURY OLSZTYN
DOI: 10.1515/pjvs-2016-0077
关键词
Adhesion; Escherichia coli; intestinal epithelial cells; pig; quantitative real-time PCR
资金
- National Natural Science Funds [31572360, 31372285]
- Genetically Modified Organisms Technology Major Project [2014ZX08006-001B]
- Science and Technology Supporting Project of Jiangsu Province [BE2015329, BE2014357]
- Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD)
An efficient and accurate method to test Escherichia coli (E. coli) adhesion to intestinal epithelial cells will contribute to the study of bacterial pathogenesis and the function of genes that encode receptors related to adhesion. This study used the quantitative real-time polymerase chain reaction (qPCR) method. qPCR primers were designed from the PILIN gene of E. coli F18ab, F18ac, and K88ac, and the pig beta-ACTIN gene. Total deoxyribonucleic acid (DNA) from E. coli and intestinal epithelial cells (IPEC-J2 cells) were used as templates for qPCR. The 2(Delta Delta Ct) formula was used to calculate the relative number of bacteria in cultures of different areas. We found that the relative numbers of F18ab, F18ac, and K88ac that adhered to IPEC-J2 cells did not differ significantly in 6-, 12-, and 24-well culture plates. This finding indicated that there was no relationship between the relative adhesion number of E. coli and the area of cells, so the method of qPCR could accurately test the relative number of E. coli. This study provided a convenient and reliable testing method for experiments involving E. coli adhesion, and also provided innovative ideas for similar detection methods.
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