Haplotype-assisted accurate non-invasive fetal whole genome recovery through maternal plasma sequencing

Abstract

BackgroundThe applications of massively parallel sequencing technology to fetal cell-free DNA (cff-DNA) have brought new insight to non-invasive prenatal diagnosis. However, most previous research based on maternal plasma sequencing has been restricted to fetal aneuploidies. To detect specific parentally inherited mutations, invasive approaches to obtain fetal DNA are the current standard in the clinic because of the experimental complexity and resource consumption of previously reported non-invasive approaches.MethodsHere, we present a simple and effective non-invasive method for accurate fetal genome recovery-assisted with parental haplotypes. The parental haplotype were firstly inferred using a combination strategy of trio and unrelated individuals. Assisted with the parental haplotype, we then employed a hidden Markov model to non-invasively recover the fetal genome through maternal plasma sequencing.ResultsUsing a sequence depth of approximately 44X against a an approximate 5.69% cff-DNA concentration, we non-invasively inferred fetal genotype and haplotype under different situations of parental heterozygosity. Our data show that 98.57%, 95.37%, and 98.45% of paternal autosome alleles, maternal autosome alleles, and maternal chromosome X in the fetal haplotypes, respectively, were recovered accurately. Additionally, we obtained efficient coverage or strong linkage of 96.65% of reported Mendelian-disorder genes and 98.90% of complex disease-associated markers.ConclusionsOur method provides a useful strategy for non-invasive whole fetal genome recovery.

Haplotype-based approach for noninvasive prenatal tests of Duchenne muscular dystrophy using cell-free fetal DNA in maternal plasma

“…In the process of decoding this HMM chain, we used the Viterbi algorithm to find the most likely sequence of hidden states. [20][21][22][23]26 Finally, we could predict the inherited haplotype and recombination breakpoints in the fetus. The detailed calculations that were used to construct the fetal genome and to determine the recombination breakpoints are available in the Supplementary Methods online.…”

“…Combining the genomic sequencing data of the parents and the fetuses, the standard fetal haplotypes could be constructed based on the trio strategy, as previously described. [20][21][22][23] Because the father has one X chromosome, we compared the accuracy of only the fetal inferred maternal SNP sties with the standard data obtained by direct sequencing. The total phased maternal SNP sites on chromosome X for each family showed a concordant ratio >99.98% (Supplementary Table S3 online).…”

“…20 To infer the fetal haplotype, we used a haplotype-based approach with a linkage relationship that was obtained from the parental haplotypes and allele frequencies that were calculated from the plasma sequencing data, as previously described. [20][21][22][23] Because the candidate mutant allele X a was inherited from the maternal chromosome X, we focused on the fetal maternalinherited chromosome X in the following analysis process. The informative SNPs that were used to infer fetal haplotype and construct the model were those maternal heterozygous SNP sites on chromosome X.…”

“…Our group recently developed a new haplotype-based strategy to recover in one step the fetal genotype and haplotype on both parentally heterozygous sites in the maternal plasma. 20 Using this approach, we successfully predicted the fetal mutation status of many autosomal recessive disorders based on the maternal plasma, including congenital adrenal hyperplasia, maple syrup urine disease, and congenital deafness. [21][22][23] However, the application of this approach to noninvasively predict the fetal mutation status in X-linked diseases has never been demonstrated.…”

Reproductive management through integration of PGD and MPS-based noninvasive prenatal screening/diagnosis for a family with GJB2-associated hearing impairment

“…SNPs, homozygous in both parents, but with different genotypes, were used for calculating fetal DNA fraction using the formula: fetal DNA fraction=2d father /(d mother +d father ), where d mother and d father stand for the allele count of the special base of the mother and the father respectively. A strategy of trios analysis based on Mendel′s law was used to deduce the parental haplotype, and a Hidden Markov Model and Viterbi algorithm were used to calculate the inherited status of maternal haplotype or paternal haplotype respectively [8,13,14].…”