90.
(11) Patent number: 2144566
(46) Date of publication: 20.01.2000
(51) Int. Cl.7: C 12 Q 1/68, C 12 N 15/00, C 12 P 19/34, A 61 K 49/00
(21) Application number: 99100157/13
(22) Application date: 05.01.1999
(71) Applicant:
Institut tsitologii i genetiki SO RAN,
NII terapii SO RAMN
(72) Inventor:
Romashchenko A.G.,
Mikhajlova S.V.,
Voevoda M.I.,
Kobzev V.F.
(73) Proprietor:
Romashchenko Aida Gerasimovna,
Mikhajlova Svetlana Vladimirovna
(98) Mail address:
630090, Novosibirsk, pr.akadem, Lavrent'eva, 10, ITsiG SO RAN, patentnyj otdel, Kuchumovoj L.Ja
(54) METHOD OF DIAGNOSIS OF PREDISPOSITION TO HEMOCHROMATOSIS
(57)
FIELD: biotechnology, molecular biology, medicine. SUBSTANCE: method involves detection of polymorphous variants of gene HLA-H (Cys282Tyr and His63Asp) based on polymorphism method of restriction fragments length. Method involves the use of primers flanking gene sites where corresponding mutations are localized and introduction of restriction endonuclease Ksp 221 site in one of primers in the case of mutation His63Asp to obtain distinct differentiation of homo- and heterozygous genotypes. Method involves amplification of DNA of gene HLA-H containing sites of possible location of mutations Cys282Tyr and His63Asp with primers 5'-CTACC-CCCAG- -AACAT-CACCA-3' and 5'-CTCCA-ATGAC-TAGGG-TGCCA-3' in the case of mutation Cys282Tyr and primers 5'-CCCTC-TCCAC-ATACC-CTTGC- -TG-3' and 5'-AAGCT-TTGGG-CTACG-TGGAT-GATCA-G- 3' in the case of mutation His63Asp followed by hydrolysis of the first of indicated amplifiers with restriction endonuclease Rsa I and the second one - with restriction endonuclease Ksp 221. Products obtained after hydrolysis of amplifiers are analyzed in polyacrylamide gel. Method ensures to screen large mass of DNA samples to detect persons showing predisposition to hemochromatosis. Diagnosis of this disease carrying out before organic damage of visceral organs ensures to treat these patients successfully and prevent the development of cirrhosis and liver cancer. EFFECT: simplified method, increased precision of diagnosis. 2 dwg, 1 ex.
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