The application of amplified TSPY and amelogenin genes from maternal plasma as a non-invasive bovine fetal DNA diagnosis

Abstract

Some studies showed analysis of fetal DNA in maternal plasma had been introduced as a new method for non-invasive prenatal diagnosis. Fetal sexing is possible at 8th week of pregnancy, using maternal blood sample testing. The aim was providing a rapid, reliable and non-invasive method for sexing of bovine fetuses. Maternal blood samples were collected from 38 pregnant cows during the 8th-38th w of gestation. Plasma was obtained by centrifugation and DNA was extracted by phenol-chloroform method from 350 􀁺L maternal plasma. Two primer pairs for bovine amelogenin, Y-encoded, and testis-specific protein gene were used to amplification three fragments; 260 bp (Y-encoded, testis-specific protein gene), 341 and 467 bp (Y and X chromosome amelogenin gene). The polymerase chain reaction has been optimized for fragments amplification. The 467 bp fragment was detected in all samples. The 341 and 260 bp fragments were detected in 24 of 38 plasma samples of cows with male fetuses. The sensitivity and specificity of test was 100% with no false negative and positive results. The results showed that phenol-chloroform method was a simple and sensational to isolation fetal DNA in maternal plasma. The polymerase chain reaction is a favorable noninvasive method for bovine fetal sexing.