Cloning and DNA Markers

Cloning a particular DNA fragment into a vector like a plasmid is a crucial step that facilitates further analysis of that sequence. Vectors from the pGEM® series offer several cloning sites suitable for fragments produced by specific restriction enzyme cuts. Fragments generated by PCR and similar TA cloning techniques can be seamlessly integrated into pGEM® T-vectors. For various subsequent applications, such as protein expression, the Flexi® Cloning System offers a convenient way to move fragments between different vectors.

Next, for typical cloning procedures you need competent cells which come in two variations: high efficiency, exceeding 10⁸cfu/μg, suitable for cloning (like JM109, HB101), and a subcloning efficiency surpassing 10⁷cfu/μg (such as HB101). An added benefit of JM109 cells is the blue/white screening capability to identify recombinants.

When aiming for protein expression, one can choose between JM109(DE3), BL21(DE3), or KRX competent cells. The Single Step (KRX) Competent Cells are tailored for effective transformation and precise protein expression control. These cells merge the top features of both processes, offering an optimized strain to assess protein expression in E. coli.