USE OF POLYMERASE CHAIN-REACTION TECHNOLOGY TO DETECT A MUTATION ASSOCIATED WITH MALIGNANT HYPERTHERMIA IN DIFFERENT PIG-TISSUES

Refinements to a novel method to detect malignant hyperthermia in swine are described. Results show that the test can be performed on whole blood and that white blood cell isolation to generate template DNA is superfluous. Storage studies demonstrate that muscle samples are best utilized within 4 days if kept at 4-degrees-C and that frozen storage at -20-degrees-C is advised for longer periods. However freeze-thaw cycles should be avoided. Fat tissue was found to be much faster than other tissues to process, particularly to run the test on a large scale. MgCl2 concentration and DNA polymerase source show small but non-significant differences in polymerase chain reaction (PCR) product formation efficiency. An internal control is proposed in order to monitor endonuclease activity and to prevent partial digestions from being erroneously interpreted. In conclusion the test is very efficient in demonstrating the mutation associated with malignant hyperthermia.