Abstract: The invention concerns a new method of detecting a rare product of a directed genetic alteration of a cultured cell. The method is applicable to any method of making the alteration provided that a pair of closely linked alterations can be made. The method consists of sequentially using allele specific polymerase chain reaction (PCR) to preferentially amplify sequences containing one of the two linked alterations coupled with a second method that detects the second change in the PCR product. The second method can be restriction digestion, traditional sequencing or pyro-sequencing. Experiments indicate that alterations as rare as one correctly altered copy in 10,000 cells can be detected.

Abstract: The invention concerns a new method of detecting a rare product of a directed genetic alteration of a cultured cell. The method is applicable to any method of making the alteration provided that a pair of closely linked alterations can be made. The method consists of sequentially using allele specific polymerase chain reaction (PCR) to preferentially amplify sequences containing one of the two linked alterations coupled with a second method that detects the second change in the PCR product. The second method can be restriction digestion, traditional sequencing or pyro-sequencing. Experiments indicate that alterations as rare as one correctly altered copy in 10,000 cells can be detected.