American Society of Hematology

Fig. 1.

$Fig. 1. Characterization of prothrombin San Antonio. / (A) The prothrombin San Antonio family. Half-filled circles: Family members identified to be heterozygous for the prothrombin San Antonio gene. Open squares: Family members identified to be homozygous for the normal allele. Prothrombin activity (numerator) and antigen (denominator) levels are indicated for several family members. P, proband; F, father; M, mother; A, Aunt; S1-S3, sons. (B) The cloned PCR product for exons 8 and 9 was analyzed by DNA sequence analysis. (asterisk) Mutation resulting in the substitution of His for Arg320. Sequences for the complementary strand of the wild-type allele (top) and the mutated prothrombin allele (bottom) were obtained directly from sequencing. Red, T; green, A; black, G; and blue, C. (C) Identification of the mutation at nucleotide 7543 in the prothrombin gene by Hha I-restriction enzyme digestion. (top) Schematic representation of the Hha I-restriction map of the PCR products for exons 8 and 9 from the wild-type (WT) and the mutant prothrombin allele. (bottom) Hha I digestion fragments of the PCR products from the proband and her family members (see part A). Fragment sizes in bp are indicated. UD, undigested. (D) Western blot analysis of the activation of prothrombin by Ecarin (in the presence of hirudin) from a normal control (N) and the maternal aunt of the proband (A). Plasma activated with Ecarin in the presence of hirudin was fractionated by 10% SDS-PAGE in the absence (top) or presence (bottom) of β-mercaptoethanol. The mobility of prothrombin is indicated (h-FII). The incubation time of plasma sample with Ecarin is indicated at the top.$

Characterization of prothrombin San Antonio.

(A) The prothrombin San Antonio family. Half-filled circles: Family members identified to be heterozygous for the prothrombin San Antonio gene. Open squares: Family members identified to be homozygous for the normal allele. Prothrombin activity (numerator) and antigen (denominator) levels are indicated for several family members. P, proband; F, father; M, mother; A, Aunt; S1-S3, sons. (B) The cloned PCR product for exons 8 and 9 was analyzed by DNA sequence analysis. (asterisk) Mutation resulting in the substitution of His for Arg320. Sequences for the complementary strand of the wild-type allele (top) and the mutated prothrombin allele (bottom) were obtained directly from sequencing. Red, T; green, A; black, G; and blue, C. (C) Identification of the mutation at nucleotide 7543 in the prothrombin gene by Hha I-restriction enzyme digestion. (top) Schematic representation of the Hha I-restriction map of the PCR products for exons 8 and 9 from the wild-type (WT) and the mutant prothrombin allele. (bottom) Hha I digestion fragments of the PCR products from the proband and her family members (see part A). Fragment sizes in bp are indicated. UD, undigested. (D) Western blot analysis of the activation of prothrombin by Ecarin (in the presence of hirudin) from a normal control (N) and the maternal aunt of the proband (A). Plasma activated with Ecarin in the presence of hirudin was fractionated by 10% SDS-PAGE in the absence (top) or presence (bottom) of β-mercaptoethanol. The mobility of prothrombin is indicated (h-FII). The incubation time of plasma sample with Ecarin is indicated at the top.

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