Ligation Independent Cloning of Polycistronic, Genetically Modified, HuMAb4D5-8 F (ab') 2, in Bacterial Plasmid
In recent years, recombinant monoclonal antibodies and their derivatives have emerged as important targeted therapy agents. Monoclonal antibodies are extremely difficult to produce. So, the cost of production is very high and many people cannot afford these drugs. In this regard, choosing inexpensiv...
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Avicenna Research Institute
2012
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| Online Access: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3558198/ |
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pubmed-35581982013-02-13 Ligation Independent Cloning of Polycistronic, Genetically Modified, HuMAb4D5-8 F (ab') 2, in Bacterial Plasmid Farahmand, Leila Majidzadeh-A, Keivan Sepehrizadeh, Zargham Mofid, Mohammad Reza Esmaeili, Rezvan Yazdi, Mojtaba Tabatabaei Original Article In recent years, recombinant monoclonal antibodies and their derivatives have emerged as important targeted therapy agents. Monoclonal antibodies are extremely difficult to produce. So, the cost of production is very high and many people cannot afford these drugs. In this regard, choosing inexpensive and easy ways to manipulate production systems such as bacterial hosts to reduce the cost of manufacturing these critical components are considered as vital step for developmental issues in recombinant expression systems. We, therefore, attempted to generate a polycistronic construct of anti HER-2 F(ab')2 fragment antibody for insertion in an expression bacterial plasmid. Also some modifications were made in the hinge region to express antibody F(ab')2 fragment in its authentic form preventing from multiple varieties of disulfide bond formation. Finally, synthesized construct was cloned in pET-32 Ek/LIC vector without using restriction enzyme digestion or ligation reactions. The results of this study showed that modified F(ab')2 fragment was simply and successfully inserted in Escherichia coli (E.coli) using the Ligation Independent Cloning technology. Avicenna Research Institute 2012 /pmc/articles/PMC3558198/ /pubmed/23407748 Text en Copyright © 2012 Avicenna Research Institute http://creativecommons.org/licenses/by-nc/3.0/ This work is licensed under a Creative Commons Attribution-NonCommercial 3.0 Unported License which allows users to read, copy, distribute and make derivative works for non-commercial purposes from the material, as long as the author of the original work is cited properly. |
| repository_type |
Open Access Journal |
| institution_category |
Foreign Institution |
| institution |
US National Center for Biotechnology Information |
| building |
NCBI PubMed |
| collection |
Online Access |
| language |
English |
| format |
Online |
| author |
Farahmand, Leila Majidzadeh-A, Keivan Sepehrizadeh, Zargham Mofid, Mohammad Reza Esmaeili, Rezvan Yazdi, Mojtaba Tabatabaei |
| spellingShingle |
Farahmand, Leila Majidzadeh-A, Keivan Sepehrizadeh, Zargham Mofid, Mohammad Reza Esmaeili, Rezvan Yazdi, Mojtaba Tabatabaei Ligation Independent Cloning of Polycistronic, Genetically Modified, HuMAb4D5-8 F (ab') 2, in Bacterial Plasmid |
| author_facet |
Farahmand, Leila Majidzadeh-A, Keivan Sepehrizadeh, Zargham Mofid, Mohammad Reza Esmaeili, Rezvan Yazdi, Mojtaba Tabatabaei |
| author_sort |
Farahmand, Leila |
| title |
Ligation Independent Cloning of Polycistronic, Genetically Modified, HuMAb4D5-8 F (ab') 2, in Bacterial Plasmid |
| title_short |
Ligation Independent Cloning of Polycistronic, Genetically Modified, HuMAb4D5-8 F (ab') 2, in Bacterial Plasmid |
| title_full |
Ligation Independent Cloning of Polycistronic, Genetically Modified, HuMAb4D5-8 F (ab') 2, in Bacterial Plasmid |
| title_fullStr |
Ligation Independent Cloning of Polycistronic, Genetically Modified, HuMAb4D5-8 F (ab') 2, in Bacterial Plasmid |
| title_full_unstemmed |
Ligation Independent Cloning of Polycistronic, Genetically Modified, HuMAb4D5-8 F (ab') 2, in Bacterial Plasmid |
| title_sort |
ligation independent cloning of polycistronic, genetically modified, humab4d5-8 f (ab') 2, in bacterial plasmid |
| description |
In recent years, recombinant monoclonal antibodies and their derivatives have emerged as important targeted therapy agents. Monoclonal antibodies are extremely difficult to produce. So, the cost of production is very high and many people cannot afford these drugs. In this regard, choosing inexpensive and easy ways to manipulate production systems such as bacterial hosts to reduce the cost of manufacturing these critical components are considered as vital step for developmental issues in recombinant expression systems. We, therefore, attempted to generate a polycistronic construct of anti HER-2 F(ab')2 fragment antibody for insertion in an expression bacterial plasmid. Also some modifications were made in the hinge region to express antibody F(ab')2 fragment in its authentic form preventing from multiple varieties of disulfide bond formation. Finally, synthesized construct was cloned in pET-32 Ek/LIC vector without using restriction enzyme digestion or ligation reactions. The results of this study showed that modified F(ab')2 fragment was simply and successfully inserted in Escherichia coli (E.coli) using the Ligation Independent Cloning technology. |
| publisher |
Avicenna Research Institute |
| publishDate |
2012 |
| url |
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3558198/ |
| _version_ |
1611950493843587072 |