Multiplex fluorescent PCR for noninvasive prenatal detection of fetal-derived paternally inherited diseases using circulatory fetal DNA in maternal plasma.
OBJECTIVE
To develop a fluorescent polymerase chain reaction (PCR) assay for the detection of circulating fetal DNA in maternal plasma and use the established multiplex in noninvasive prenatal genetic diagnosis and its further applications in forensic casework.


METHODS
The DNA template was extracted from 47 pregnant women and the whole blood samples from the stated biological fathers were used to detect genotype. Using multiplex fluorescent PCR at 16 different polymorphic short tandem repeat (STR) loci, maternal DNA extracted from plasma samples at early pregnancy, medium pregnancy and late pregnancy were used to detect genotype. Their husbands' DNA was also used for fetal genotype ascertainment.


RESULTS
Multiplex fluorescent PCR with 16 polymorphic short tandem repeats revealed the presence of fetal DNA in all cases. Every pregnant women/husband pair was informative in at least 3 of 16 loci. The chances of detecting paternally inherited fetal alleles ranged from 66.67 to 94.12%. They are 66.67% in early pregnancy, 85.71% in medium pregnancy and 94.12% in late pregnancy. The accuracy of Multiplex PCR assay to detect fetal DNA was 100%.


CONCLUSIONS
Circulating fetal DNA analysis can be used as a possible alternative tool in routine laboratory prenatal diagnosis in the near future; this highly polymorphic STR multiplex has greatly improved the chances of detecting paternally inherited fetal alleles compared with other fetal DNA detection systems that use fetus-derived Y sequences to detect only male fetal DNA in maternal plasma. Our proposed technique can be applied to both female and male fetuses, which provides a sensitive, accurate and efficient method for noninvasive prenatal genetic diagnosis and forensic casework.
Tang
Dong-ling
DL
Department of Clinical Laboratory, Renmin Hospital of Wuhan University, Hubei Province, PR China.
Li
Yan
Y
Zhou
Xin
X
Li
Xia
X
Zheng
Fang
F
D000483:Alleles; D004247:DNA / Q000097:blood*; D005220:Fathers; D005260:Female; D005333:Fetus / Q000378:metabolism*; D030342:Genetic Diseases, Inborn / Q000097:blood / Q000175:diagnosis* / Q000235:genetics; D005838:Genotype; D006801:Humans; D008297:Male; D018895:Microsatellite Repeats / Q000235:genetics; D009035:Mothers; D016133:Polymerase Chain Reaction / Q000379:methods*; D011247:Pregnancy; D011296:Prenatal Diagnosis / Q000379:methods*
D004247:DNA