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Background  Myosin-binding protein C3  A31P  ( MYBPC3-A31P ) missense mutation is a genetic deviation associated with the development of hypertrophic cardiomyopathy (HCM) in Maine Coon cats. The standard detection of the  MYBPC3-A31P  mutation is complicated, time-consuming, and expensive. Currently, there has been a focus on the speed and reliability of diagnostic tools. Therefore, this study aimed to develop a loop-mediated isothermal amplification assay (LAMP) coupled with a lateral flow dipstick (LFD) test to detect  MYBPC3-A31P  mutations in Maine Coon cats.    Materials and Methods  Fifty-five Maine Coon cats were enrolled in this study, and blood samples were collected.  MYBPC3-A31P  was genotyped by DNA sequencing. Primers for LAMP with a LFD test were designed. The optimal conditions were determined, including temperature and time to completion for the reaction. The sensitivity of  A31P -LAMP detection was compared between agarose gel electrophoresis (the standard method) and the LFD test. The  A31P -LAMP-LFD test was randomly performed on seven cats (four with the  A31P  mutation and three wild-type cats).    Results  The  A31P -LAMP procedure was able to distinguish between cats with  MYBPC3-A31P  wild-type cats and  MYBPC3-A31P  mutant cats. The LAMP reactions were able to be completed in 60 min at a single temperature of 64◦C. Moreover, this study demonstrated that  A31P -LAMP coupled with the LFD test allowed for  A31P  genotype detection at a lower DNA concentration than agarose gel electrophoresis.    Discussions  This new  A31P -LAMP with a LFD test is a successful and reliable assay with a rapid method, cost-effectiveness, and low requirements for sophisticated equipment for the detection of  MYBPC3-A31P  mutations. Thus, this assay has excellent potential and can be recognized as a novel screening test for hypertrophic cardiomyopathy associated with  MYBPC3-A31P  mutations in felines.

frontiers in veterinary science

Development of a Loop-Mediated Isothermal Amplification Assay Coupled With a Lateral Flow Dipstick Test for Detection of Myosin Binding Protein C3 A31P Mutation in Maine Coon Cats