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Biochemistry and Molecular Biology Education
Article . 2004 . Peer-reviewed
License: Wiley Online Library User Agreement
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Gene amplification by PCR and subcloning into a GFP‐fusion plasmid expression vector as a molecular biology laboratory course*

Authors: Amy B. Mulnix; Michael Deibel; Joshua A. Bornhorst; Joshua A. Bornhorst;

Gene amplification by PCR and subcloning into a GFP‐fusion plasmid expression vector as a molecular biology laboratory course*

Abstract

AbstractA novel experimental sequence for the advanced undergraduate laboratory course has been developed at Earlham College. Utilizing recent improvements in molecular techniques for a time‐sensitive environment, undergraduates were able to create a chimera of a selected gene and green fluorescent protein (GFP) in a bacterial expression plasmid over the course of a single semester in a weekly 3‐h laboratory period. Students designed PCR primers for amplification of the selected gene using computational DNA sequence analysis tools. During the experimental portion of the course, students amplified and ligated the target DNA into a commercially available GFP expression vector. Following transformation of the ligation product, plasmids were harvested from the resulting bacterial colonies and were analyzed by restriction digestion to confirm the creation of the chimeric GFP‐DNA. This course gave students valuable experience with commonly used molecular techniques in an authentic research project. In addition, students gained experience with experimental design and execution. The techniques presented here are flexible and can be generalized for use with almost any DNA sequence and expression vector. This series also serves as an example of how faculty can adapt their ongoing research projects to the undergraduate laboratory.

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citations
This is an alternative to the "Influence" indicator, which also reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
BIP!Citations provided by BIP!
popularity
This indicator reflects the "current" impact/attention (the "hype") of an article in the research community at large, based on the underlying citation network.
BIP!Popularity provided by BIP!
influence
This indicator reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
BIP!Influence provided by BIP!
impulse
This indicator reflects the initial momentum of an article directly after its publication, based on the underlying citation network.
BIP!Impulse provided by BIP!
7
Average
Average
Average
bronze