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Isolation and Puri cation of CNF1 (Cytotoxic necro- sis factor 1) produced from bacteria Escherichia coli and study its role against apoptosis in vitro | ||
| Iraqi Journal of Cancer and Medical Genetics | ||
| Article 1, Volume 8, Issue 2, 2015, Pages 157-163 | ||
| Authors | ||
| Nahi Y. Yaseen; May T. Flayih; Muthanna H. Hassan | ||
| Abstract | ||
| Through the period April - December 2012, urine specimens were collected from186 patiants attending ve hospitals. Sev- enty six isolates of uropathogenic E. coli were obtained from these samples. These isolates were identi ed as E. coli ac- cording to the cultural and morphological characteristics on MacConkey agar, eosin methylene blue agar, biochemical test (vi- tek 2 system), and API 20 E system. The results showed that only four E. coli isolates (E4, E8, E23, E36) produced cytotoxic necrosis factor 1(CNF1), and E23 was the more ef cient isolate in CNF1 production and have high activity on tissue culture. CNF1 produced from this isolate was puri ed by two steps; the rst was by ion exchange chromatography using DEAE- Sephadex, and the second step was gel ltration by Sephacryl S-100. Also study Polymerase Chain Reaction (PCR) for cnf1 gene and used primer cnf1. The size of ampli ed gene was 543pb. The cytotoxic effect of different concentrations of CNF1 on cancer lines (RD, AMN3, , Hela, PC3 and HePG2) at 72 hr were examined .The effect was manifested multinucleated and enlargement in cell line. The low IC50 was 104μg/ml against HePG2cancer cells after 72 hr. Also study multiparameter cytotoxic assay. The results revealed low concentration of cytochrome c, total nuclear intensity, cell membrane permeability in comparative positive control. The mitochondria concentration was high compared to the positive control on of cell lines in the concentration (100,25, 12.5,6.25). | ||
| Keywords | ||
| CNF; Escherichia coli; apoptosis; In Vitro | ||
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