PURIFICATION OF TOXIN FROM E. COLI ISOLATE Pe68 BY ION-EXCHANGE CHROMATOGRAPHY
Volume
3, Issue 1
Pages:
440-446
Year of Publication:
April, 2017
Journal of Applied Science and Engineering Methodologies
ISSN:
2395–5341
| Citation: Mary Conice and M.S.Usha."PURIFICATION OF TOXIN FROM E. COLI ISOLATE Pe68 BY ION-EXCHANGE CHROMATOGRAPHY" Journal of Applied Science and Engineering Methodologies,Vol.3,No.1(2017):440-446.
BibTex
@article{chand2017aderiv, author = { Mary Conice,M.S.Usha}, title = {PURIFICATION OF TOXIN FROM E. COLI ISOLATE Pe68 BY ION-EXCHANGE CHROMATOGRAPHY}, journal = {Journal of Applied Science and Engineering Methodologies}, issue_date = {25}, volume = {3}, number = {1}, month = {Apr}, year = {2017}, issn = {2395–5341}, pages = {440-446}, numpages = {7}, url = {http://www.jasem.in/2017/31440446.html}, publisher = {Journal of Applied Science and Engineering Methodologies}, address = {Chennai, India} } |
| DOI: | Full Text Download |
Abstract:
E. coli isolate pe68 which showed positive to hlyA toxin was subjected to protein purification by Ion-exchange chromatography. The purification steps involed salting out, dialysis and ion-exchange separation carried out in column packed with ion-exchanger. In the present study Sephrose 6B as fast-flow weak anion and Carboxymethyl Cellulose as fast-flow weak cation was used as stationary phase and elution buffer was used as mobile phase. Different concentrations of Tris HCl and Sodium Chloride solution i.e., 1M of sodium chloride solution and 1M Tris HCl buffer with pH 7 was used as elution buffer. Different fractions of elution were collected every 5 minutes and were subjected to protein estimation by Lowry’s method. Protein obtained from above purification steps were subjected to molecular weight determination by SDS-PAGE using low-protein molecular weight markers. SDS-PAGE of the protein sample resulted in a single band close to the protein marker with 33 kDa.
Keywords:Escherichia coli, ion-exchanger chromatography, SDS-PAGE.
E. coli isolate pe68 which showed positive to hlyA toxin was subjected to protein purification by Ion-exchange chromatography. The purification steps involed salting out, dialysis and ion-exchange separation carried out in column packed with ion-exchanger. In the present study Sephrose 6B as fast-flow weak anion and Carboxymethyl Cellulose as fast-flow weak cation was used as stationary phase and elution buffer was used as mobile phase. Different concentrations of Tris HCl and Sodium Chloride solution i.e., 1M of sodium chloride solution and 1M Tris HCl buffer with pH 7 was used as elution buffer. Different fractions of elution were collected every 5 minutes and were subjected to protein estimation by Lowry’s method. Protein obtained from above purification steps were subjected to molecular weight determination by SDS-PAGE using low-protein molecular weight markers. SDS-PAGE of the protein sample resulted in a single band close to the protein marker with 33 kDa.
Keywords:Escherichia coli, ion-exchanger chromatography, SDS-PAGE.
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