Massively parallel sequencing has been reported only to apply in noninvasive prenatal diagnosis of trisomy 21, 18 and 13 based on cell-free fetal DNA, due to it is limited amount of fragmented fetal DNA 
. This also makes noninvasive MPS-based prenatal diagnosis difficult to detect all the chromosomal aneuploidies and the sexual chromosome abnormality accurately. In this study, we demonstrated for the first time that combined MPS with powerful bioinformatics analysis method can accurately diagnosis fetal aneuploidy and imbalanced chromosomal structural abnormalities. In fact, this study reports the first retrospective use of MPS (so called next generation sequencing) for prenatal diagnostics of chromosomal imbalance rearrangements to date and shows that it is practically feasible on a large-scale prenatal diagnosis of fetal chromosomal abnormalities.
By establishing a normal control sequencing tag data set, we have been able to demonstrate this new approach only requires a minimum among of DNA materials (100 ng) to achieve the identification of aneuploidies with a ultra low sequencing coverage (0.01×). Comparing to the golden standard (G-Band karyotyping) in clinical practice, MPS has no time limitation. Also as long as 100 ng genomic DNA can be extracted from tissues at any gestational weeks, MPS can be performed and report all fetal aneuploidies in 7 days. If necessary, more sequence reads can be performed to detect whether microdeletion or microduplication exists in the fetal genome which may result in severe developmental retardation. In our study, tissues obtained from fetus, such as amniotic fluid, CVS and placenta can be analyzed without cell culture since 100 ng genomic DNA is sufficient for library preparation and sequencing. So it can be used for the research into the molecular mechanism of miscarriage, stillbirth and fetal death when tissues are difficult to culture. Furthermore, our validation study on the 5 arrayCGH samples show that when more sophisticated sequencing protocols and bioinformatics algorithms are applied to the analysis, it is possible to detect smaller size chromosomal copy number variations as well as complex rearrangements across the whole genome of the fetus, such as balanced chromosomal arrangements,structure variations or even single-gene disorders. With the application of third-generation sequencing system in clinical laboratories, such as Miseq/Illumina and Ion Torrent PGM/Life Technologies, the whole process will take less time and acceptable price. Thus, it is likely that MPS will play an increasingly important role in the future development of prenatal screening and diagnosis.
A potential weakness of the study was that in the figure of describing the ratio of unique reads in each chromosome, chromosome19 and 22 have a differently huge coefficient of variation because of their extremely high GC content and made the detection of trisomy19 and trisomy22 difficult. Further study will be set up to deal with these problems, for example the computational correction of GC content among chromosomes. Other chromosomal abnormalities, such as balanced translocation or incomplete aneuploidy caused by mosaics or partial duplication or deletion of a chromosome should, in principle, also be detectable. Further studies are required to determine the effectiveness of massively parallel genomic sequencing in detecting these rare aberrations. Another weakness for the new methodology was the starting materials for library construction; with the conventional Illumina library construction approach100 ng genomic DNA of fetus was required. To further reduce the risk to pregnancies it would be important to reduce the amount for AF or CVS samples, other library construction methods such as using in vitro transposition may be an alternative solution.
In conclusion, we have demonstrated the usefulness of massively parallel sequencing to detect fetal aneuploidy and imbalanced chromosomal abnormalities of genomic DNA in prenatal samples. In principle, massively parallel sequencing can also reveal other features of the genomic material from amniotic fluid such as histone modifications as well as epigenetic DNA methylation. With the rapid reduction of sequencing cost, we expect that the strategy described in this article will become a powerful tool in the detection of all kinds of chromosomal abnormalities in clinical settings.