LaFORA'S DISEASE GENE

Abstract

A novel gene (EPM2A) that is deleted or mutated in people with Lafora's disease is described. The EPM2A gene encodes a protein having an active catalytic site of a protein tyrosine phosphatase. Many different sequence mutations as well as several microdeletions in EPM2A have been found that co-segregate with Lafora's disease.

What is claimed is: 
     
         1 . A method of detecting Lafora's disease in a mammal comprising detecting the presence or absence of a mutation in a nucleic acid molecule containing a sequence which is associated with Lafora's disease, wherein the mutation is selected from the group consisting of a point mutation, an insertion of one or more nucleotides and a deletion of one or more nucleotides in the nucleic acid molecule as compared to the nucleic acid sequence as shown in SEQ ID NO:1 and wherein the presence of a mutation indicates that the mammal has Lafora's disease or is a carrier of Lafora's disease. 
     
     
         2 . The method according to  claim 1 , comprising detecting a mutation or deletion in a region of the nucleic sequence between markers DS61003 and DS61042 as shown in  FIGS. 1A-1B . 
     
     
         3 . The method according to  claim 1 , wherein the mutation is detected by a method comprising:
 (a) amplifying the nucleic acid sequence containing a sequence which is associated with Lafora's disease with primers in a polymerase chain reaction;   (b) digesting the amplified sequences with a restriction endonuclease; and   (c) determining the size of the digested sequence wherein a difference in size as compared to the size of a digested sequence from SEQ ID NO:1 that has been amplified and digested according to steps (a) and (b) indicates that the mammal has Lafora's disease or is a carrier of Lafora's disease.   
     
     
         4 . The method according to  claim 1 , wherein the point mutation is a missense mutation or a nonsense mutation. 
     
     
         5 . The method according to  claim 4 , wherein the nonsense mutation comprises the replacement of nucleotide C at position 721 in SEQ ID NO:1 with nucleotide T. 
     
     
         6 . A method according to  claim 5 , wherein the C to T change is detected by a method comprising:
 (a) amplifying the nucleic acid sequences in the sample with primers H1F (5′-GAATGCTCTTTCCACTTTGC-3) (SEQ ID NO:7) and PTPR (5′-GGCTCCTTAGGGAAATCAG-3′) (SEQ ID NO:8) in a polymerase chain reaction;   (b) digesting the amplified sequences with the restriction endonuclease HaeIII; and   (c) determining the size of the digested sequences wherein the presence of a fragment of approximately 199 bp indicates that the sample is from an animal with Lafora's disease or an animal that is a carrier of Lafora's disease.   
     
     
         7 . The method according to  claim 4 , wherein the missense mutation comprises the replacement of nucleotide G at position 836 in SEQ ID NO:1 with nucleotide A. 
     
     
         8 . A method according to  claim 7 , wherein the G to A change is detected by a method comprising:
 (a) amplifying the nucleic acid sequences in the sample with primers H1F (5′-GAATGCTCTTTCCACTTTGC-3) (SEQ ID NO:7) and PTPR (5′-GGCTCCTTAGGGAAATCAG-3′) (SEQ ID NO:8) in a polymerase chain reaction;   (b) digesting the amplified sequences with the restriction endonuclease Pstl; and   (c) determining the size of the digested sequences wherein the presence of at least one fragment of approximately 520 bp indicates that the sample is from an animal that does not have Lafora's disease or an animal that is a carrier of Lafora's disease.   
     
     
         9 . The method according to  claim 4 , wherein the nonsense mutation comprises the replacement of nucleotide C at position 163 in SEQ ID NO:1 with nucleotide T. 
     
     
         10 . The method according to  claim 4 , wherein the missense mutation comprises the replacement of nucleotide A at position 146 in SEQ ID NO:1 with nucleotide G. 
     
     
         11 . The method according to  claim 4 , wherein the nonsense mutation comprises the replacement of nucleotide G at position 412 in SEQ ID NO:1 with nucleotide T. 
     
     
         12 . The method according to  claim 4 , wherein the missense mutation comprises the replacement of nucleotide A at position 878 in SEQ ID NO:1 with nucleotide T. 
     
     
         13 . The method according to  claim 4 , wherein the nonsense mutation comprises the replacement of nucleotide G at position 179 in SEQ ID NO:1 with nucleotide A. 
     
     
         14 . The method according to  claim 4 , wherein the missense mutation comprises the replacement of nucleotide C at position 322 in SEQ ID NO:1 with nucleotide T. 
     
     
         15 . The method according to  claim 4 , wherein the missense mutation comprises the replacement of nucleotide T at position 94 in SEQ ID NO:1 with nucleotide G. 
     
     
         16 . The method according to  claim 1 , wherein the mutation is an insertion of nucleotide A at position 800 in SEQ ID NO:1. 
     
     
         17 . The method according to  claim 1 , wherein the mutation is a deletion mutation. 
     
     
         18 . The method according to  claim 17 , wherein nucleotide G at position 235 in SEQ ID NO:1 is deleted. 
     
     
         19 . The method according to  claim 17 , wherein there is a deletion of exons 1 and 2. 
     
     
         20 . The method according to  claim 17 , wherein there is a deletion of exon 2. 
     
     
         21 . A method of according to  claim 20 , comprising detecting a deletion of about 25 kb or 75 kb in the sequence shown in SEQ ID NO:1, and further:
 (a) amplifying the nucleic acid sequences in the sample with primers JRGXBF (5′-TCCATTGTGCTAATGCTATCTC-3′) (SEQ ID NO:9) and JRGXBR (5′-TCAGCTTGCTTTGAGGATATTT-3′) (SEQ ID NO:10) in a polymerase chain reaction; and   (b) detecting amplified sequence wherein the absence of an amplified sequence indicates that the sample is from an animal with Lafora's disease.