BACKGROUNDThe embryonic genome, including genotypes and haplotypes, contains all the information for preimplantation genetic diagnosis, representing great potential for mendelian disorder carriers to conceive healthy babies.METHODSWe developed a strategy to obtain the full embryonic genome for a β-thalassemia-carrier couple to have a healthy second baby. We carried out sequencing for single blastomere cells and the family trio and further developed the analysis pipeline, including recovery of the missing alleles, removal of the majority of errors, and phasing of the embryonic genome.RESULTSThe final accuracy for homozygous and heterozygous single-nucleotide polymorphisms reached 99.62% and 98.39%, respectively. The aneuploidies of embryos were detected as well. Based on the comprehensive embryonic genome, we effectively performed whole-genome mendelian disorder diagnosis and human leukocyte antigen matching tests.CONCLUSIONSThis retrospective study in a β-thalassemia family demonstrates a method for embryo genome recovery through single-cell sequencing, which permits detection of genetic variations in preimplantation genetic diagnosis. It shows the potential of single-cell sequencing technology in preimplantation genetic diagnosis clinical practices.

Purpose: This study demonstrates noninvasive prenatal testing (NIPT) for Duchenne muscular dystrophy (DMD) using a newly developed haplotype-based approach. Methods: Eight families at risk for DMD were recruited for this study. Parental haplotypes were constructed using target-region sequencing data from the parents and the probands. Fetal haplotypes were constructed using a hidden Markov model through maternal plasma DNA sequencing. The presence of haplotypes linked to the maternal mutant alleles in males indicated affected fetuses. This method was further validated by comparing the inferred single-nucleotide polymorphism (SNP) genotypes to the direct sequencing results of fetal genomic DNA. Prenatal diagnosis was confirmed with amniocentesis, and those results were interpreted in a blinded fashion. Results: The results showed an average accuracy of 99.98% for the total inferred maternal SNPs. With a mean depth of 30× achieved in the 10-Mb target region of each sample, the noninvasive results were consistent with those of the invasive procedure. Conclusion: This is the first report of NIPT for DMD and the first application of a haplotype-based approach in NIPT for X-linked diseases. With further improvements in accuracy, this haplotype-based strategy could be feasible for NIPT for DMD and even other X-linked single-gene disorders. Genet Med 17 11, 889–896.

Concave-disdyakis triacontahedral palladium nanocrystals (C-DTH Pd NCs) bound with 120 {631} high index facets were prepared by electrochemically shape-controlled method in deep eutectic solvent (DES). It has found that both the adsorption of urea derived from EDS and the upper (En) and lower (EL) limit potentials of the square-wave potential applied in the synthesis are contributed synergetically in controlling the shape of Pd NCs. The formation of C-DTH Pd NCs with well-defined shape was achieved by the dynamic interaction between urea adsorption at Eu and growth at EL. In situ FTIR spectroscopic studies revealed that the urea adsorbates at Eu play a crucial role in shape evolution, especially in the formation of C-DTH Pd NCs. It has demonstrated that the as synthesized C-DTH Pd NCs enclosed by {631} high-index facets exhibit higher electrocatalytic activity than Pd NCs with other shapes bound by {111} low-index facets (octahedral (OH) and icosahedral (IH)) toward ethanol electrooxidation in alkaline medium.

Prenatal diagnosis of congenital adrenal hyperplasia (CAH) is of clinical significance because in utero treatment is available to prevent virilization of an affected female fetus. However, traditional prenatal diagnosis of CAH relies on genetic testing of fetal genomic DNA obtained using amniocentesis or chorionic villus sampling, which is associated with an increased risk of miscarriage. The aim of this study was to demonstrate the feasibility of a new haplotype-based approach for the noninvasive prenatal testing of CAH due to 21-hydroxylase deficiency. Parental haplotypes were constructed using target-region sequencing data of the parents and the proband. With the assistance of the parental haplotypes, we recovered fetal haplotypes using a hidden Markov model (HMM) through maternal plasma DNA sequencing. In the genomic region around the CYP21A2 gene, the fetus inherited the paternal haplotype ‘0’ alleles linked to the mutant CYP21A2 gene, but the maternal haplotype ‘1’ alleles linked to the wild-type gene. The fetus was predicted to be an unaffected carrier of CAH, which was confirmed by genetic analysis of fetal genomic DNA from amniotic fluid cells. This method was further validated by comparing the inferred SNP genotypes with the direct sequencing data of fetal genomic DNA. The result showed an accuracy of 96.41% for the inferred maternal alleles and an accuracy of 97.81% for the inferred paternal alleles. The haplotype-based approach is feasible for noninvasive prenatal testing of CAH.

Three-dimensional (3D) aggregates composed of 5.0 +/- 0.4 nm palladium nanocrystallites (PdNCs) were prepared using ultrasonics in the palladium (II) chloride (PdCl2) H2O/EtOH (5 : 1, V/V) solution by the addition of a quantity of Poly(vinyl pyrrolidone) (PVP) and hexadecyl trimethyl ammonium bromide (CTAB). The morphologies, crystal structures and the optical properties of 3D Pd nanostructures were characterised by X-ay diffraction, X-ray photoelectron spectroscopy, transmission electron microscopy, selected area electron diffraction and UV-visible absorption spectroscopy, respectively. The mechanism of sonochemical reduction of Pd (II) ions was also discussed. The results showed that the formation of aggregates of PdNCs depended on the interaction between PVP and CTAB. Moreover, the electrocatalytic properties of 3D Pd nanostructures modified glassy carbon electrode for ethanol oxidation had been also investigated by cyclic voltammetry. The material exhibited remarkable electrocatalytic behaviour for ethanol oxidation in 1M KOH and appeared as a promising candidate for use in direct ethanol fuel cells.

Purpose: The goals of our study were to develop a noninvasive prenatal test for autosomal recessive monogenic conditions and to prove its overall feasibility and potential for clinical integration. Methods: We recruited a pregnant woman and her spouse, who had a proband child suffering from congenital deafness, and obtained the target-region sequencing data from a semicustom array that used genomic and maternal plasma DNA from three generations of this family. A haplotype-assisted strategy was developed to detect whether the fetus inherited the pathogenic mutations in the causative gene, GJB2 . The parental haplotype was constructed using a trio strategy through two different processes, namely, the grandparent-assisted haplotype phasing process and the proband-assisted haplotype phasing process. The fetal haplotype was deduced afterward based on both the maternal plasma sequencing data and the parental haplotype. Results: The accuracy levels of paternal and maternal haplotypes obtained by grandparent-assisted haplotype phasing were 99.01 and 97.36%, respectively, and the proband-assisted haplotype phasing process yielded slightly lower accuracies of 98.73 and 96.79%, respectively. Fetal inheritance of the pathogenic gene was deduced correctly in both processes. Conclusion: Our study indicates that the strategy of haplotype-based noninvasive prenatal testing for monogenic conditions has potential applications in clinical practice. Genet Med 16 12, 972–976.

Objective: To develop a fetal sex determination method based on maternal plasma sequencing (MPS), assess its performance and potential use in X-linked disorder counseling.Methods: 900 cases of MPS data from a previous study were reviewed, in which 100 and 800 cases were used as training and validation set, respectively. The percentage of uniquely mapped sequencing reads on Y chromosome was calculated and used to classify male and female cases. Eight pregnant women who are carriers of Duchenne muscular dystrophy (DMD) mutations were recruited, whose plasma were subjected to multiplex sequencing and fetal sex determination analysis.Results: In the training set, a sensitivity of 96% and false positive rate of 0% for male cases detection were reached in our method. The blinded validation results showed 421 in 423 male cases and 374 in 377 female cases were successfully identified, revealing sensitivity and specificity of 99.53% and 99.20% for fetal sex determination, at as early as 12 gestational weeks. Fetal sex for all eight DMD genetic counseling cases were correctly identified, which were confirmed by amniocentesis.Conclusions: Based on MPS, high accuracy of non-invasive fetal sex determination can be achieved. This method can potentially be used for prenatal genetic counseling.

Background: Next generation sequencing (NGS) is now being used for detecting chromosomal abnormalities in blastocyst trophectoderm (TE) cells from in vitro fertilized embryos. However, few data are available regarding the clinical outcome, which provides vital reference for further application of the methodology. Here, we present a clinical evaluation of NGS-based preimplantation genetic diagnosis/screening (PGD/PGS) compared with single nucleotide polymorphism (SNP) array-based PGD/PGS as a control.Results: A total of 395 couples participated. They were carriers of either translocation or inversion mutations, or were patients with recurrent miscarriage and/or advanced maternal age. A total of 1,512 blastocysts were biopsied on D5 after fertilization, with 1,058 blastocysts set aside for SNP array testing and 454 blastocysts for NGS testing. In the NGS cycles group, the implantation, clinical pregnancy and miscarriage rates were 52.6% (60/114), 61.3% (49/80) and 14.3% (7/49), respectively. In the SNP array cycles group, the implantation, clinical pregnancy and miscarriage rates were 47.6% (139/292), 56.7% (115/203) and 14.8% (17/115), respectively. The outcome measures of both the NGS and SNP array cycles were the same with insignificant differences. There were 150 blastocysts that underwent both NGS and SNP array analysis, of which seven blastocysts were found with inconsistent signals. All other signals obtained from NGS analysis were confirmed to be accurate by validation with qPCR. The relative copy number of mitochondrial DNA (mtDNA) for each blastocyst that underwent NGS testing was evaluated, and a significant difference was found between the copy number of mtDNA for the euploid and the chromosomally abnormal blastocysts. So far, out of 42 ongoing pregnancies, 24 babies were born in NGS cycles; all of these babies are healthy and free of any developmental problems.Conclusions: This study provides the first evaluation of the clinical outcomes of NGS-based pre-implantation genetic diagnosis/screening, and shows the reliability of this method in a clinical and array-based laboratory setting. NGS provides an accurate approach to detect embryonic imbalanced segmental rearrangements, to avoid the potential risks of false signals from SNP array in this study.

Carbon supported Pt nanocatalysts(Pt/C) are widely used in fuel cells. In this work, three kinds of Pt/C catalysts were successfully prepared via a chemical reduction method by using carbon black pretreated with different methods, i.e., HNO3-pretreated carbon(Pt/C-HNO3), H2O2-pretreated carbon(Pt/C-H2O2) and unpretreated carbon(Pt/C). The morphologies and properties of the as-prepared Pt/C catalysts were characterized by SEM, HR-TEM, CV and COad stripping. Pt nanoparticles were dispersed well in these three Pt/C catalysts with average particle size of 3.4, 3.9 and 4.5 nm respectively. HR-TEM observation indicated that Pt/C-HNO3 possessed high-density of step atoms and COad stripping experiments showed that Pt/C-HNO3 exhibited the best anti-poisoning property. Comparing the catalytic activities of three as-prepared Pt/C catalysts toward ethanol electrooxidation with commercial Pt/C catalyst(Pt/C JM) showed that the as-prepared catalysts exhibited higher activity and stability than those of Pt/C JM and the order was Pt/C-HNO3Pt/C-H2O2Pt/CPt/C JM. Especially for Pt/C-HNO3, its activity and stability were 1.5 times and 1.9 times of those of Pt/C JM respectively.

ABSTRACT Objective The objective of this study is to assess the performance of noninvasive prenatal testing for trisomies 21 and 18 on the basis of massively parallel sequencing of cell‐free DNA from maternal plasma in twin pregnancies. Method A double‐blind study was performed over 12 months. A total of 189 pregnant women carrying twins were recruited from seven hospitals. Maternal plasma DNA sequencing was performed to detect trisomies 21 and 18. The fetal karyotype was used as gold standard to estimate the sensitivity and specificity of sequencing‐based noninvasive prenatal test. Results There were nine cases of trisomy 21 and two cases of trisomy 18 confirmed by karyotyping. Plasma DNA sequencing correctly identified nine cases of trisomy 21 and one case of trisomy 18. The discordant case of trisomy 18 was an unusual case of monozygotic twin with discordant fetal karyotype (one normal and the other trisomy 18). The sensitivity and specificity of maternal plasma DNA sequencing for fetal trisomy 21 were both 100% and for fetal trisomy 18 were 50% and 100%, respectively. Conclusion Our study further supported that sequencing‐based noninvasive prenatal testing of trisomy 21 in twin pregnancies could be achieved with a high accuracy, which could effectively avoid almost 95% of invasive prenatal diagnosis procedures. © 2013 John Wiley & Sons, Ltd.

Background Copy number variations (CNVs) represent an important type of genetic variation that deeply impact phenotypic polymorphisms and human diseases. The advent of high-throughput sequencing technologies provides an opportunity to revolutionize the discovery of CNVs and to explore their relationship with diseases. However, most of the existing methods depend on sequencing depth and show instability with low sequence coverage. In this study, using low coverage whole-genome sequencing (LCS) we have developed an effective population-scale CNV calling (PSCC) method. Methodology/Principal Findings In our novel method, two-step correction was used to remove biases caused by local GC content and complex genomic characteristics. We chose a binary segmentation method to locate CNV segments and designed combined statistics tests to ensure the stable performance of the false positive control. The simulation data showed that our PSCC method could achieve 99.7%/100% and 98.6%/100% sensitivity and specificity for over 300 kb CNV calling in the condition of LCS (∼2×) and ultra LCS (∼0.2×), respectively. Finally, we applied this novel method to analyze 34 clinical samples with an average of 2× LCS. In the final results, all the 31 pathogenic CNVs identified by aCGH were successfully detected. In addition, the performance comparison revealed that our method had significant advantages over existing methods using ultra LCS. Conclusions/Significance Our study showed that PSCC can sensitively and reliably detect CNVs using low coverage or even ultra-low coverage data through population-scale sequencing.