US 4,395,486 A · Method for the direct analysis of sickle cell anemia

Abstract

A direct diagnosis of sickle cell anemia by a restriction endonuclease assay with an enzyme which recognizes the nucleotide base sequence CTNAG (where N is any base), such as Dde I, for the sickle cell allele (.beta..sup.s gene) through molecular hybridization. Following enzyme cleavage, the resulting DNA restriction fragments are separated by molecular weight and transferred to filter paper. A probe is utilized for hybridization that is complementary to the 5' end of the .beta. globin gene. The banding pattern of individuals with normal hemoglobin shows two bands (approximately 175 bp and 201 bp), sickle cell trait individuals exhibit an additional band (approximately 376 bp) and individuals with sickle cell anemia show the band at approximately 376 bp with a concommitant loss of the band at approximately 175 bp. Prenatal and postnatal diagnosis of sickle cell anemia is possible with the present method.

Claims27

Dependent claims

A method for the direct analysis of sickle cell anemia in the human .beta. gene, comprising the steps of:

(a) obtaining an effective amount of amniotic fluid cells from a human;
(b) isolating DNA from said cells;
(c) digesting said DNA with restriction enzyme Dde I to form DNA fragments;
(d) separating said DNA fragments according to their respective molecular weights to form a pattern for said fragments on filter paper;
and
(e) detecting the absence of an approximate 175 base pair fragment and the presence of an approximate 376 base pair fragment on said pattern, thereby indicating the sickle cell genotype.

3 more elements

Dependencies are inferred from the claim text.

Method for the direct analysis of sickle cell anemia

Life of this patent

Method for the direct analysis of sickle cell anemia

Life of this patent

Abstract

Claims27