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231226s2022 xx |||||o 00| ||eng c |
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|a 10.2144/btn-2022-0085
|2 doi
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|a (NLM)36398840
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|a DE-627
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|e rakwb
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|a eng
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| 100 |
1 |
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|a Yang, Zhibo
|e verfasserin
|4 aut
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| 245 |
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|a A simple and economical site-directed mutagenesis method for large plasmids by direct transformation of two overlapping PCR fragments
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|c 2022
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|a Text
|b txt
|2 rdacontent
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|a ƒaComputermedien
|b c
|2 rdamedia
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|a ƒa Online-Ressource
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|2 rdacarrier
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|a Date Completed 23.11.2022
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|a Date Revised 04.12.2022
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|a published: Print-Electronic
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|a Citation Status MEDLINE
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|a Despite the development of various methods and commercial kits, site-directed mutagenesis of large plasmids remains a challenge in many laboratories. A site-directed mutagenesis method was developed for large plasmids by directly transforming two overlapping PCR fragments into Escherichia coli. This method successfully generated mutations for plasmids of 8.3 kb and 11.0 kb with high efficiencies. The method only requires Q5 DNA polymerase and DpnI, which greatly reduces costs. The procedure is simple, including PCR reaction, DpnI treatment and transformation. This simple, efficient and economical site-directed mutagenesis method for large plasmids is likely to be widely applied in the future
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|a Journal Article
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|a Research Support, Non-U.S. Gov't
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|a Escherichia coli
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|a Gibson assembly
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|a PCR-based site-directed mutagenesis
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|a Q5 DNA polymerase
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|a Q5 site-directed mutagenesis
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|a QuikChange site-directed mutagenesis
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|a in vivo assembly
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|a in vivo recombination
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|a overlapping extension PCR
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| 650 |
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|a phusion site-directed mutagenesis
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|a DNA-Directed DNA Polymerase
|2 NLM
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| 650 |
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|a EC 2.7.7.7
|2 NLM
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| 700 |
1 |
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|a Chen, Zan
|e verfasserin
|4 aut
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| 700 |
1 |
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|a Zhang, Yueping
|e verfasserin
|4 aut
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| 773 |
0 |
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|i Enthalten in
|t BioTechniques
|d 1993
|g 73(2022), 5 vom: 15. Nov., Seite 239-245
|w (DE-627)NLM012627046
|x 1940-9818
|7 nnas
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| 773 |
1 |
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|g volume:73
|g year:2022
|g number:5
|g day:15
|g month:11
|g pages:239-245
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|u http://dx.doi.org/10.2144/btn-2022-0085
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