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Using this site-directed mutagenesis technique, three variants of the mouse major histocompatibility complex class I gene have been generated, cloned and analyzed. Screening of mutant clones showed at least 98% mutagenesis efficiency. All clones sequenced contained the desired mutations, and a low frequency of random substitution estimated to occur at approx. 1 in 4000 nt was detected. This method represents a significant improvement over standard site-directed mutagenesis because it is faster, simpler, and nearly 100% efficient in generating mutant products.