Application of a target array Comparative Genomic Hybridization to prenatal diagnosis

BMC Medical Genetics RAespeaprchliacrtaiclteion of a target array Comparative Genomic Hybridization to prenatal diagnosis Ji Hyeon Park Jung Hoon Woo 0 Sung Han Shim Song-Ju Yang 0 Young Min Choi Kap-Seok Yang 0 Dong Hyun Cha 1 0 Department of Bio Chip Service, Macrogen Inc. , Seoul , Korea 1 Department of Obstetrics and Gynecology, CHA Gangnam Medical Center, CHA University , Seoul , Republic of Korea, Seoul , Korea Background: While conventional G-banded karyotyping still remains a gold standard in prenatal genetic diagnoses, the widespread adoption of array Comparative Genomic Hybridization (array CGH) technology for postnatal genetic diagnoses has led to increasing interest in the use of this same technology for prenatal diagnosis. We have investigated the value of our own designed DNA chip as a prenatal diagnostic tool for detecting submicroscopic deletions/ duplications and chromosome aneuploidies. Methods: We designed a target bacterial artificial chromosome (BAC)-based aCGH platform (MacArray M-chip), which specifically targets submicroscopic deletions/duplications for 26 known genetic syndromes of medical significance observed prenatally. To validate the DNA chip, we obtained genomic DNA from 132 reference materials generated from patients with 22 genetic diseases and 94 clinical amniocentesis samples obtained for karyotyping. Results: In the 132 reference materials, all known genomic alterations were successfully identified. In the 94 clinical samples that were also subjected to conventional karyotyping, three cases of balanced chromosomal aberrations were not detected by aCGH. However, we identified eight cases of microdeletions in the Yq11.23 chromosomal region that were not found by conventional karyotyping. This region harbors the DAZ gene, and deletions may lead to nonobstructive spermatogenesis. Conclusions: We have successfully designed and applied a BAC-based aCGH platform for prenatal diagnosis. This platform can be used in conjunction with conventional karyotyping and will provide rapid and accurate diagnoses for the targeted genomic regions while eliminating the need to interpret clinically-uncertain genomic regions. - Background Speed and precision are two major requirements in prenatal chromosome analyses. Conventional G-banded karyotyping remains the gold standard in prenatal genetic diagnosis, but it is time-consuming and labor-intensive. Routinely, about 10-14 days is required to obtain the result and this may increase the patient's anxiety. To overcome these limitations, rapid fluorescent in situ hybridization (FISH), quantitative fluorescent polymerase chain reaction (QF-PCR), and multiplex ligation-dependent probe amplification (MLPA) have been developed and are widely used as adjuncts to conventional methods for detecting common chromosome aneuploidies such as trisomies 21, 13, 18, X and Y [1-6]. However, only a few loci may be tested at a time, so all those methods can usually be performed only in a limited manner based on phenotype. In addition, conventional G-banded karyotyping can hardly detect small deletions and duplications that result in serious clinical conditions such as congenital anomalies, mental retardation and developmental delay during the fetal stage as well as after birth. Recently, array comparative genomic hybridization (aCGH) has been used in prenatal as well as postnatal clinical cytogenetics to detect submicroscopic chromosomal imbalances [7-9]. This technique gives rapid results and multiplex detection of both numerical and unbalanced structural abnormalities with much higher resolution and wider coverage than conventional karyotyping and other molecular cytogenetic techniques. Several types of platform with different resolutions for aCGH have been developed. More recently, genome-wide oligonucleotide aCGH chips with several-kb resolutions have become commercially available. However, for clinical purposes, especially prenatal diagnosis, these high-resolution whole-genome aCGH chips are not adequate because of difficulties of interpretation mainly resulting from benign copy number variations (CNVs), hybridization quality and cost. On the other hand, targeted bacterial artificial chromosome (BAC)-based aCGH methods have been successfully used in prenatal diagnosis [7,10-12]. In this study, we have developed and evaluated a low density BAC-array prenatal DNA chip targeted to 26 known genetic syndromes of medical significance, caused by 19 microdeletions/duplications as well as numerical changes in chromosomes 13, 18, 21, X and Y. Methods Subjects To validate the array, because we could not obtain four microdeletion syndromes - Sotos syndrome, monosomy 1p36, SRY region of Yp and Kallmann syndrome - we used 132 reference materials of 22 genetic disorders, 15 for microdeletions or duplications and 7 for chromosome aneuploidies, as positive controls. The cell lines were purchased from the Coriell Cell Resource http://ccr.coriell.org/nigms/products/pdr.ht (...truncated)