title: Reliable detection of paternal SNPs within deletion breakpoints for non-invasive prenatal exclusion of homozygous α-thalassemia in maternal plasma.
abstract: Reliable detection of large deletions from cell-free fetal DNA (cffDNA) in maternal plasma is challenging, especially when both parents have the same deletion owing to a lack of specific markers for fetal genotyping. In order to evaluate the efficacy of a non-invasive prenatal diagnosis (NIPD) test to exclude α-thalassemia major that uses SNPs linked to the normal paternal α-globin allele, we established a novel protocol to reliably detect paternal SNPs within the (--(SEA)) breakpoints and performed evaluation of the diagnostic potential of the protocol in a total of 67 pregnancies, in whom plasma samples were collected prior to invasive obstetrics procedures in southern China. A group of nine SNPs identified within the deletion breakpoints were scanned to select the informative SNPs in each of the 67 couples DNA by multiplex PCR based mini-sequencing technique. The paternally inherited SNP allele from cffDNA was detected by allele specific real-time PCR. A protocol for reliable detection of paternal SNPs within the (--(SEA)) breakpoints was established and evaluation of the diagnostic potential of the protocol was performed in a total of 67 pregnancies. In 97% of the couples one or more different SNPs within the deletion breakpoint occurred between paternal and maternal alleles. Homozygosity for the (--(SEA)) deletion was accurately excluded in 33 out of 67 (49.3%, 95% CI, 25.4-78.6%) pregnancies through the implementation of the protocol. Protocol was completely concordant with the traditional reference methods, except for two cases that exhibited uncertain results due to sample hemolysis. This method could be used as a routine NIPD test to exclude gross fetal deletions in α-thalassemia major, and could further be employed to test for other diseases due to gene deletion.
authors: Department of Medical Genetics, School of Basic Medical Sciences, Southern Medical University, Guangzhou, Guangdong, People's Republic of China.
authors: Ti-Zhen
authors: TZ
authors: Yan
authors: Qiu-Hua
authors: QH
authors: Mo
authors: Ren
authors: R
authors: Cai
authors: Xue
authors: X
authors: Chen
authors: Cui-Mei
authors: CM
authors: Zhang
authors: Yan-Hui
authors: YH
authors: Liu
authors: Ya-Jun
authors: YJ
authors: Chen
authors: Wan-Jun
authors: WJ
authors: Zhou
authors: Fu
authors: F
authors: Xiong
authors: Xiang-Min
authors: XM
authors: Xu
mesh_terms: D000328:Adult; D000483:Alleles; D002681:China; D004247:DNA / Q000097:blood; D005260:Female; D017353:Gene Deletion; D005787:Gene Frequency; D006579:Heterozygote; D006801:Humans; D008297:Male; D009035:Mothers; D010949:Plasma / Q000378:metabolism*; D016133:Polymerase Chain Reaction / Q000379:methods; D020641:Polymorphism, Single Nucleotide; D011247:Pregnancy; D011296:Prenatal Diagnosis / Q000379:methods*; D012680:Sensitivity and Specificity; D017085:alpha-Thalassemia / Q000175:diagnosis / Q000235:genetics*
chemical_list: D004247:DNA