Title: Introduction to Recombinant DNA
Course: BTEC-1100-301-302-303-F21 > Section: Module 4 - Applied Molecular Biology

In module 3 we amplified and purified or gene of interest, bglA from B. Halodurans.  In this module we will clone our gene into a plasmid vector called pLATE31 to create recombinant DNA.  Molecular cloning allows us to bring together different sources of DNA to produce novel sequences.  In our case we are combining the pLATE DNA and our bglA DNA to produce a beta-galactosidase protein for the enzymatic digestion of cellulose.   Recombinant DNA was first achieved in the early 1970s using the enzymatic activity of restriction enzymes and DNA ligase to first cut and then paste DNA from an organism into a plasmid.

Click here for a larger version of this graphic. CC BY-SA 3.0: Kelvinsong

For our project we will be using a more modern method of cloning that eliminates the need for restriction enzymes as well as DNA ligase.  The method we will use is called ligation independent cloning or LIC.

Learning Objectives:

- Clone a gene using an ligation independent cloning (LIC) reaction.
- Transform bacteria using the results of LIC.
- Screen bacteria for the presence of the plasmid and gene using gel electrophoresis.
- Isolate plasmid DNA from positive clones.
- Analyze sequencing data.