TY - JOUR
T1 - Polymerase chain reaction-restriction fragment length polymorphism method for identifying carriers of hemophilia A in Japanese brown cattle
AU - Khalaj, Maryam
AU - Abbasi, Abdol Rahim
AU - Tsuji, Takehito
AU - Moritomo, Yasuo
AU - Shimojo, Kenichi
AU - Kunieda, Tetsuo
PY - 2006/2/1
Y1 - 2006/2/1
N2 - Hemophilia A is a severe congenital bleeding disorder characterized by subcutaneous hematoma and hemorrhage into muscles resulting from a deficiency of blood coagulation factor VIII. The authors have recently reported two cases of hemophilia A in Japanese Brown cattle and identified a nucleotide substitution in the factor VIII gene, resulting in an amino acid substitution of Leu to His, as a possible cause of the deficiency. In the present study, a simple and effective polymerase chain reaction (PCR)-based diagnostic method was developed to identify carriers of this disorder, using a mismatch primer in combination with restriction enzyme digestion. The PCR reaction amplified a 118 bp fragment, which was not digested by the BspT104I restriction enzyme in affected animals but was digested into two fragments in normal animals. Both digested and undigested fragments were observed in carrier animals. This method was applied to identify the carriers of hemophilia A in a population of Japanese Brown cattle. By screening 155 DNA samples from Japanese Brown cattle, except for the dam of the two probands, no carriers were identified. It was therefore concluded that the probands represent isolated cases of hemophilia A, and that the frequency of the mutant allele in the Japanese Brown cattle population is very low.
AB - Hemophilia A is a severe congenital bleeding disorder characterized by subcutaneous hematoma and hemorrhage into muscles resulting from a deficiency of blood coagulation factor VIII. The authors have recently reported two cases of hemophilia A in Japanese Brown cattle and identified a nucleotide substitution in the factor VIII gene, resulting in an amino acid substitution of Leu to His, as a possible cause of the deficiency. In the present study, a simple and effective polymerase chain reaction (PCR)-based diagnostic method was developed to identify carriers of this disorder, using a mismatch primer in combination with restriction enzyme digestion. The PCR reaction amplified a 118 bp fragment, which was not digested by the BspT104I restriction enzyme in affected animals but was digested into two fragments in normal animals. Both digested and undigested fragments were observed in carrier animals. This method was applied to identify the carriers of hemophilia A in a population of Japanese Brown cattle. By screening 155 DNA samples from Japanese Brown cattle, except for the dam of the two probands, no carriers were identified. It was therefore concluded that the probands represent isolated cases of hemophilia A, and that the frequency of the mutant allele in the Japanese Brown cattle population is very low.
KW - Bleeding disorder
KW - Factor VIII
KW - Hemophilia A
KW - Japanese brown cattle
KW - Polymerase chain reaction-restriction fragment length polymorphism
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U2 - 10.1111/j.1740-0929.2006.00329.x
DO - 10.1111/j.1740-0929.2006.00329.x
M3 - Article
AN - SCOPUS:33644984218
VL - 77
SP - 122
EP - 125
JO - Animal Science Journal
JF - Animal Science Journal
SN - 1344-3941
IS - 1
ER -