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2011•Biotechnology(Faisalabad)Requires access

A Modified and Rapid Protocol for the Transformation of Plasmid DNA

Wang You-ru

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Abstract

Objective:The factors effecting on the transformation frequency of E.coli DH5α and JM109 by plasmid DNA were studied.A rapid protocol for the transformation of plasmid DNA using CaCl2 was reported.Method:5-10min incubation of cells with DNA on ice,heated-shock at 42℃ for 60s,and then 2min incubation of cells with DNA on ice,followed by incubation of cells at 37℃ for 2min and 8 min of shaking at 37℃.Spreading onto agar plates prewarmed to 37℃,resulting in more than 105 colonies/μg plasmid DNA.Result:The manipulation time was shorened from 2 hours to 20 minutes.Conclusion:This protocol is simpler and less time-wasted than the normal one.

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What this paper is about

Objective:The factors effecting on the transformation frequency of E.coli DH5α and JM109 by plasmid DNA were studied.A rapid protocol for the transformation of plasmid DNA using CaCl2 was reported.Method:5-10min incubation of cells with DNA on ice,heated-shock at 42℃ for 60s,and then 2min incubation of cells with DNA on ice,followed by incubation of cells at 37℃ for 2min and 8 min of shaking at 37℃.Spreading onto agar plates prewarmed to 37℃,resulting in more than 105 colonies/μg plasmid DNA.Result:The manipulation time was shorened from 2 hours to 20 minutes.Conclusion:This protocol is simpler and less time-wasted than the normal one.

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Available abstract

Objective:The factors effecting on the transformation frequency of E.coli DH5α and JM109 by plasmid DNA were studied.A rapid protocol for the transformation of plasmid DNA using CaCl2 was reported.Method:5-10min incubation of cells with DNA on ice,heated-shock at 42℃ for 60s,and then 2min incubation of cells with DNA on ice,followed by incubation of cells at 37℃ for 2min and 8 min of shaking at 37℃.Spreading onto agar plates prewarmed to 37℃,resulting in more than 105 colonies/μg plasmid DNA.Result:The manipulation time was shorened from 2 hours to 20 minutes.Conclusion:This protocol is simpler and less time-wasted than the normal one.

Key concepts: Plasmid, Transformation (genetics), Incubation, DNA, Molecular biology, Plasmid preparation, Incubation period, Agar

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