Combined Overlap Extension PCR Method for Improved Site Directed Mutagenesis
The combined overlap extension PCR (COE-PCR) method developed in this work combines the strengths of the overlap extension PCR (OE-PCR) method with the speed and ease of the asymmetrical overlap extension (AOE-PCR) method. This combined method allows up to 6 base pairs to be mutated at a time and re...
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| creator | Hussain, Mohd Hasnain Md Chong, Nikson Fatt-Ming |
| description | The combined overlap extension PCR (COE-PCR) method developed in this work combines the strengths of the overlap extension PCR (OE-PCR) method with the speed and ease of the asymmetrical overlap extension (AOE-PCR) method. This combined method allows up to 6 base pairs to be mutated at a time and requires a total of 40–45 PCR cycles. A total of eight mutagenesis experiments were successfully carried out, with each experiment mutating between two to six base pairs. Up to four adjacent codons were changed in a single experiment. This method is especially useful for codon optimization, where doublet or triplet rare codons can be changed using a single mutagenic primer set, in a single experiment. |
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This combined method allows up to 6 base pairs to be mutated at a time and requires a total of 40–45 PCR cycles. A total of eight mutagenesis experiments were successfully carried out, with each experiment mutating between two to six base pairs. Up to four adjacent codons were changed in a single experiment. This method is especially useful for codon optimization, where doublet or triplet rare codons can be changed using a single mutagenic primer set, in a single experiment.</description><identifier>ISSN: 2314-6133</identifier><identifier>EISSN: 2314-6141</identifier><identifier>DOI: 10.1155/2016/8041532</identifier><identifier>PMID: 27995143</identifier><language>eng</language><publisher>Cairo, Egypt: Hindawi Publishing Corporation</publisher><subject>Base Pair Mismatch - genetics ; Codon ; Codon - genetics ; Deoxyribonucleic acid ; DNA ; DNA - genetics ; E coli ; Enzymes ; Genes ; Methods ; Multiplex Polymerase Chain Reaction - methods ; Mutagenesis ; Mutagenesis, Site-Directed - methods ; Mutation ; Mutation - genetics ; Protein expression</subject><ispartof>BioMed research international, 2016-01, Vol.2016 (2016), p.1-7</ispartof><rights>Copyright © 2016 Hasnain Hussain and Nikson Fatt-Ming Chong.</rights><rights>COPYRIGHT 2016 John Wiley & Sons, Inc.</rights><rights>Copyright © 2016 Hasnain Hussain and Nikson Fatt-Ming Chong. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.</rights><rights>Copyright © 2016 H. Hussain and N. F.-M. Chong. 2016</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c532t-3ef8af60d610cc5ef95cbfabde09034b6e73a4108de29f2c07d154e4ae3479c3</citedby><cites>FETCH-LOGICAL-c532t-3ef8af60d610cc5ef95cbfabde09034b6e73a4108de29f2c07d154e4ae3479c3</cites><orcidid>0000-0002-2630-450X</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC5138438/pdf/$$EPDF$$P50$$Gpubmedcentral$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC5138438/$$EHTML$$P50$$Gpubmedcentral$$Hfree_for_read</linktohtml><link.rule.ids>230,314,723,776,780,881,27901,27902,53766,53768</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/27995143$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><contributor>Ongecha, John M.</contributor><creatorcontrib>Hussain, Mohd Hasnain Md</creatorcontrib><creatorcontrib>Chong, Nikson Fatt-Ming</creatorcontrib><title>Combined Overlap Extension PCR Method for Improved Site Directed Mutagenesis</title><title>BioMed research international</title><addtitle>Biomed Res Int</addtitle><description>The combined overlap extension PCR (COE-PCR) method developed in this work combines the strengths of the overlap extension PCR (OE-PCR) method with the speed and ease of the asymmetrical overlap extension (AOE-PCR) method. 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This method is especially useful for codon optimization, where doublet or triplet rare codons can be changed using a single mutagenic primer set, in a single experiment.</description><subject>Base Pair Mismatch - genetics</subject><subject>Codon</subject><subject>Codon - genetics</subject><subject>Deoxyribonucleic acid</subject><subject>DNA</subject><subject>DNA - genetics</subject><subject>E coli</subject><subject>Enzymes</subject><subject>Genes</subject><subject>Methods</subject><subject>Multiplex Polymerase Chain Reaction - methods</subject><subject>Mutagenesis</subject><subject>Mutagenesis, Site-Directed - methods</subject><subject>Mutation</subject><subject>Mutation - genetics</subject><subject>Protein expression</subject><issn>2314-6133</issn><issn>2314-6141</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2016</creationdate><recordtype>article</recordtype><sourceid>RHX</sourceid><sourceid>EIF</sourceid><sourceid>BENPR</sourceid><recordid>eNqN0c9rFDEUB_BBFFtqb55lwItg1-b3JBehrFULWyrae8hkXnZTZpI1mVntf2-WXZfVU3NJQj58eS-vql5j9AFjzi8JwuJSIoY5Jc-qU0IxmwnM8PPDmdKT6jznB1SWxAIp8bI6IY1SHDN6Wi3mcWh9gK6-20Dqzbq-_j1CyD6G-tv8e30L4yp2tYupvhnWKW6K_OFHqD_5BHYst9tpNEsIkH1-Vb1wps9wvt_PqvvP1_fzr7PF3Zeb-dViZkuZ44yCk8YJ1AmMrOXgFLetM20HSCHKWgENNQwj2QFRjljUdJgzYAYoa5SlZ9XHXex6agfoLIQxmV6vkx9MetTReP3vS_ArvYwbzTGVjMoS8G4fkOLPCfKoB58t9L0JEKessRSSUsYa8gTKMVFKMFXo2__oQ5xSKB9RFCui9HaklqYH7YOLpUS7DdVXnKAG84aKoi52yqaYcwJ36A4jvZ283k5e7ydf-JvjHzngv3Mu4P0OrHzozC__xDgoBpw50koqgugfC2e9rg</recordid><startdate>20160101</startdate><enddate>20160101</enddate><creator>Hussain, Mohd Hasnain Md</creator><creator>Chong, Nikson Fatt-Ming</creator><general>Hindawi Publishing Corporation</general><general>John Wiley & Sons, Inc</general><general>Hindawi Limited</general><scope>ADJCN</scope><scope>AHFXO</scope><scope>RHU</scope><scope>RHW</scope><scope>RHX</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>3V.</scope><scope>7QL</scope><scope>7QO</scope><scope>7T7</scope><scope>7TK</scope><scope>7U7</scope><scope>7U9</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8FD</scope><scope>8FE</scope><scope>8FG</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABUWG</scope><scope>AFKRA</scope><scope>ARAPS</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BGLVJ</scope><scope>BHPHI</scope><scope>C1K</scope><scope>CCPQU</scope><scope>CWDGH</scope><scope>DWQXO</scope><scope>FR3</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>H94</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>LK8</scope><scope>M0S</scope><scope>M1P</scope><scope>M7N</scope><scope>M7P</scope><scope>P5Z</scope><scope>P62</scope><scope>P64</scope><scope>PIMPY</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PRINS</scope><scope>7X8</scope><scope>5PM</scope><orcidid>https://orcid.org/0000-0002-2630-450X</orcidid></search><sort><creationdate>20160101</creationdate><title>Combined Overlap Extension PCR Method for Improved Site Directed Mutagenesis</title><author>Hussain, Mohd Hasnain Md ; Chong, Nikson Fatt-Ming</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c532t-3ef8af60d610cc5ef95cbfabde09034b6e73a4108de29f2c07d154e4ae3479c3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2016</creationdate><topic>Base Pair Mismatch - genetics</topic><topic>Codon</topic><topic>Codon - genetics</topic><topic>Deoxyribonucleic acid</topic><topic>DNA</topic><topic>DNA - genetics</topic><topic>E coli</topic><topic>Enzymes</topic><topic>Genes</topic><topic>Methods</topic><topic>Multiplex Polymerase Chain Reaction - methods</topic><topic>Mutagenesis</topic><topic>Mutagenesis, Site-Directed - methods</topic><topic>Mutation</topic><topic>Mutation - genetics</topic><topic>Protein expression</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Hussain, Mohd Hasnain Md</creatorcontrib><creatorcontrib>Chong, Nikson Fatt-Ming</creatorcontrib><collection>الدوريات العلمية والإحصائية - e-Marefa Academic and Statistical Periodicals</collection><collection>معرفة - المحتوى العربي الأكاديمي المتكامل - e-Marefa Academic Complete</collection><collection>Hindawi Publishing Complete</collection><collection>Hindawi Publishing Subscription Journals</collection><collection>Hindawi Publishing Open Access</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>ProQuest Central (Corporate)</collection><collection>Bacteriology Abstracts (Microbiology B)</collection><collection>Biotechnology Research Abstracts</collection><collection>Industrial and Applied Microbiology Abstracts (Microbiology A)</collection><collection>Neurosciences Abstracts</collection><collection>Toxicology Abstracts</collection><collection>Virology and AIDS Abstracts</collection><collection>Health & Medical Collection</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>Medical Database (Alumni Edition)</collection><collection>Technology Research Database</collection><collection>ProQuest SciTech Collection</collection><collection>ProQuest Technology Collection</collection><collection>ProQuest Natural Science Collection</collection><collection>Hospital Premium Collection</collection><collection>Hospital Premium Collection (Alumni Edition)</collection><collection>ProQuest Central (Alumni) (purchase pre-March 2016)</collection><collection>ProQuest Central (Alumni Edition)</collection><collection>ProQuest Central UK/Ireland</collection><collection>Advanced Technologies & Aerospace Collection</collection><collection>ProQuest Central Essentials</collection><collection>Biological Science Collection</collection><collection>ProQuest Central</collection><collection>Technology Collection</collection><collection>Natural Science Collection</collection><collection>Environmental Sciences and Pollution Management</collection><collection>ProQuest One Community College</collection><collection>Middle East & Africa Database</collection><collection>ProQuest Central Korea</collection><collection>Engineering Research Database</collection><collection>Health Research Premium Collection</collection><collection>Health Research Premium Collection (Alumni)</collection><collection>ProQuest Central Student</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>SciTech Premium Collection</collection><collection>ProQuest Health & Medical Complete (Alumni)</collection><collection>ProQuest Biological Science Collection</collection><collection>Health & Medical Collection (Alumni Edition)</collection><collection>Medical Database</collection><collection>Algology Mycology and Protozoology Abstracts (Microbiology C)</collection><collection>Biological Science Database</collection><collection>Advanced Technologies & Aerospace Database</collection><collection>ProQuest Advanced Technologies & Aerospace Collection</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Publicly Available Content Database</collection><collection>ProQuest One Academic Eastern Edition (DO NOT USE)</collection><collection>ProQuest One Academic</collection><collection>ProQuest One Academic UKI Edition</collection><collection>ProQuest Central China</collection><collection>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>BioMed research international</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Hussain, Mohd Hasnain Md</au><au>Chong, Nikson Fatt-Ming</au><au>Ongecha, John M.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Combined Overlap Extension PCR Method for Improved Site Directed Mutagenesis</atitle><jtitle>BioMed research international</jtitle><addtitle>Biomed Res Int</addtitle><date>2016-01-01</date><risdate>2016</risdate><volume>2016</volume><issue>2016</issue><spage>1</spage><epage>7</epage><pages>1-7</pages><issn>2314-6133</issn><eissn>2314-6141</eissn><abstract>The combined overlap extension PCR (COE-PCR) method developed in this work combines the strengths of the overlap extension PCR (OE-PCR) method with the speed and ease of the asymmetrical overlap extension (AOE-PCR) method. 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| source | MEDLINE; Wiley Online Library Open Access; PubMed Central; Alma/SFX Local Collection; PubMed Central Open Access |
| subjects | Base Pair Mismatch - genetics Codon Codon - genetics Deoxyribonucleic acid DNA DNA - genetics E coli Enzymes Genes Methods Multiplex Polymerase Chain Reaction - methods Mutagenesis Mutagenesis, Site-Directed - methods Mutation Mutation - genetics Protein expression |
| title | Combined Overlap Extension PCR Method for Improved Site Directed Mutagenesis |
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