Inverse Pcr Mutagenesis

This procedure utilizes the basic principle of IPCR to introduce a genetic change within a plasmid. As in traditional IPCR, the primers are oriented ''back-to-back'' but anneal in opposite directions around the plasmid, and one of the primers contain the required mutation. PCR of the plasmid with a proofreading enzyme will introduce the genetic change present in one primer and yield a blunt-ended product. Self-ligation of the final product will reproduce the original plasmid plus the introduced mutation (Fig. 3). This procedure can be utilized for point mutations or insertions. If a region of the plasmid is to be deleted, primers can be designed on either side of this area, and the subsequent PCR product when religated will be complete except for the region that the primers flanked (Fig. 4).

Transformation of ligated mutated plasmids into Escherichia coli gives a high efficiency of mutant plasmid isolation (approximately 80%).

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Getting Started With Dumbbells

Getting Started With Dumbbells

The use of dumbbells gives you a much more comprehensive strengthening effect because the workout engages your stabilizer muscles, in addition to the muscle you may be pin-pointing. Without all of the belts and artificial stabilizers of a machine, you also engage your core muscles, which are your body's natural stabilizers.

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