Use of single nucleotide polymorphism microarrays to distinguish between balanced and normal chromosomes in embryos from a translocation carrier

Transcription

1 CASE REPORT Use of single nucleotide polymorphism microarrays to distinguish between balanced and normal chromosomes in embryos from a translocation carrier Nathan R. Treff, Ph.D., a,b Xin Tao, M.S., a Wendy J. Schillings, M.D., c Paul A. Bergh, M.D., a,b Richard T. Scott Jr., M.D., a,b and Brynn Levy, Ph.D. a,d a Reproductive Medicine Associates of New Jersey, Morristown, New Jersey; b Division of Reproductive Endocrinology and Infertility, Department of Obstetrics Gynecology and Reproductive Science, University of Medicine and Dentistry of New Jersey (UMDNJ) Robert Wood Johnson Medical School, New Brunswick, New Jersey; c Reproductive Medicine Associates of Pennsylvania, Allentown, Pennsylvania; and d Department of Pathology, College of Physicians and Surgeons, Columbia University, New York, New York Objective: To prove the ability to distinguish between balanced and normal chromosomes in embryos from a translocation carrier. Design: Case report. Setting: Academic center for reproductive medicine. Patient(s): Woman with a balanced translocation causing Alagille syndrome seeking preimplantation genetic diagnosis (PGD). Intervention(s): Blastocyst biopsy for PGD. Main Outcome Measure(s): Consistency of 3 methods of embryo genetic analysis (real-time polymerase chain reaction, single nucleotide polymorphism [SNP] microarray, and fluorescence in situ hybridization [FISH]) and normalcy in the newborn derived from PGD. Result(s): PGD was applied to 48 embryos. Real-time polymerase chain reaction, SNP microarray, and FISH demonstrated 100% consistency, although FISH failed to detect aneuploidies observed by comprehensive SNP microarray-based analyses. Two blastocysts were identified to be normal for all 3 factors using SNP microarray technology alone. The 2 normal embryos were transferred back to the patient, resulting in the delivery of a healthy boy with a normal karyotype. Conclusion(s): This is the first report of validation and successful clinical application of microarray-based PGD to distinguish between balanced and normal chromosomes in embryos from a translocation carrier. (Fertil Steril Ò 2011;96:e Ó2011 by American Society for Reproductive Medicine.) Key Words: Aneuploidy, preimplantation embryo, Alagille syndrome, translocation, microdeletion Received February 23, 2011; revised April 12, 2011; accepted April 13, 2011; published online May 14, N.R.T. has received research grants from EMD Serono, Ferring, Schering Plough, and Barr Pharmaceuticals. X.T. has nothing to disclose. W.J.S. has nothing to disclose. P.A.B. has nothing to disclose. R.T.S. has received research grants from EMD Serono, Ferring, Schering Plough, and Barr Pharmaceuticals. B.L. is a member of the advisory board for Affymetrix. Reprint requests: Nathan R. Treff, Ph.D., Reproductive Medicine Associates of New Jersey Research, 111 Madison Avenue, Suite 100, Morristown, NJ ( ntreff@rmanj.com). It is estimated that 1 in 625 individuals carries a balanced translocation (1). In most cases, carriers are phenotypically normal, but they are at increased risk for clinical pregnancy loss and offspring with congenital abnormalities and/or mental retardation as a result of unbalanced segregation during gametogenesis. A clinical phenotype may also result if there is disruption of critical gene(s) at the breakpoint regions. Studies using newer molecular cytogenetic techniques have now demonstrated that microdeletions are responsible for a proportion of cases in which an apparently balanced translocation is associated with phenotypic abnormality (2 4). In fact, small genomic imbalances such as microdeletions, as well as duplications or inversions, occur at or near the chromosomal breakpoint in 10% to 60% of cases (4 12). A good example is Alagille syndrome (ALGS), which is caused by mutations in the Jagged1 gene (13, 14). There are now a few reports of patients with ALGS with apparently balanced translocations, all of which share a common breakpoint on chromosome 20 at p12.2 (13 15). A microdeletion encompassing the Jagged1 gene has been demonstrated in some of these cases (13, 14) and presumed in the others. Individuals carrying this type of genetic abnormality who are interested in producing healthy offspring through preimplantation genetic diagnosis (PGD) would require more careful screening than is currently available. The primary aim of PGD would be to distinguish between embryos carrying the balanced translocation (and accompanying microdeletion) and those with truly normal chromosomes. A previously developed single nucleotide polymorphism (SNP) microarray based method of aneuploidy screening (16) e58 Fertility and Sterility â Vol. 96, No. 1, July /$36.00 Copyright ª2011 American Society for Reproductive Medicine, Published by Elsevier Inc. doi: /j.fertnstert

2 FIGURE 1 Workup results of a patient with ALGS. (A) Conventional karyotype analysis confirmed an apparently balanced translocation, 46,XX,t(2;20)(q21;p12.2). (B) SNP oligonucleotide microarray analysis copy number (CN) analysis of chromosome 20 revealed a microdeletion (including the Jagged1 locus) that causes ALGS. The expanded view of cytoband 20p12.2 indicates the dbsnp rs identifiers for each of the 6 SNPs identified as informative and used in the subsequent triple-factor PGD of embryos derived from the patient during 2 IVF cycles. (C) Real-time polymerase chain reaction results for lymphocyte samples from the patient, partner, or mixture of the 2 illustrating the expected genotypes of homozygous opposite for each intended parent and heterozygous for the mixtures. Genomic DNA of each sample served as a control for the expected genotypes of the 5-cell samples. (D) Confirmation of the 2.36-Mb deletion using fluorescence in situ hybridization (FISH) with probe RPC I10 (Jagged1 locus). Only 1 copy of the Jagged1 locus, on the normal chromosome 20, is present. Fertility and Sterility â e59

3 TABLE 1 Preclinical ALGS real-time PCR assay validation. Sample type Analysis Informative SNPs rs rs rs rs rs rs Maternal gdna Microarray result C T G T A A gdna TaqMan result C T G T A A 5-cell C T G T A A 5-cell C T G T A A 5-cell C T G T A A 5-cell C T G T A A Paternal gdna Microarray result TT CC AA CC GG GG gdna TaqMan result TT CC AA CC GG GG 5-cell TT CC AA CC GG GG 5-cell TT CC AA CC GG GG 5-cell TT CC AA CC GG GG 5-cell TT CC AA CC GG GG Mixed parental 4 maternal:2 paternal cells TaqMan result CT TC GA TC AG AG 4 maternal:2 paternal cells CT TC GA TC AG AG 4 maternal:2 paternal gdna CT TC GA TC AG AG 4 maternal:2 paternal gdna CT TC GA TC AG AG Note: PCR ¼ polymerase chain reaction. offers an opportunity do so. This method was recently used to simultaneously detect unbalanced chromosome derivatives and aneuploidy of all 24 chromosomes in embryos from patients carrying balanced translocations (17). Therefore, the ability of SNP microarrays to detect an inherited microdeletion associated with an apparently balanced translocation was investigated in the present study. This proof-of-principle study also represents the first clinical application of simultaneous detection of a single gene disorder, unbalanced chromosome derivatives, and aneuploidy of all 24 chromosomes from the same embryo biopsy. MATERIALS AND METHODS Patient and Case Workup A 28-year-old patient with ALGS with an apparently balanced translocation 46,XX,t(2;20)(q21;p12.2) (Fig. 1A) presented for PGD. The presence of a putative microdeletion was investigated by analysis of DNA isolated from peripheral blood of the patient and her partner. Chromosome copy number was evaluated by SNP microarray analysis as previously described (16). Informative SNPs were identified based on the criteria that the patient and partner were homozygous for 2 different (opposite) alleles within the putative microdeletion. SNP microarray data described in this study were deposited in the Gene Expression Omnibus of the National Center for Biotechnology Information and are accessible through GEO series accession number GSE21732 ( geo). Assays TaqMan SNP genotyping based predesigned allelic discrimination assays (Applied Biosystems Inc., Foster City, CA) were selected from the informative SNPs on the Affymetrix Nsp I SNP microarray (Affymetrix, Santa Clara, CA) that defined the boundaries of the microdeletion in the patient (Table 1). Each assay was evaluated on lymphocyte samples derived from the patient and her partner either separately (5 cells) or in a 2:1 mixture (patient cell:partner cell ratio; 6 cells) to mimic heterozygosity. Cells were prepared by alkaline lysis as previously described (16). Preamplification of the SNP assay target DNA was conducted on each sample using TaqMan PreAmp Master Mix as recommended by the supplier (Applied Biosystems Inc.) and before conventional allelic discrimination based analysis. Embryos found suitable for cryopreservation in the first of 2 IVF treatment cycles were biopsied for trophectoderm tissue at the blastocyst stage as previously described (18). Half of each trophectoderm biopsy was preincubated in a hypotonic solution and fixed using 3:1 methanol:acetic acid solution. Three rounds of fluorescence in situ hybridization (FISH) were performed using probes for the unbalanced translocation derivatives, the microdeletion, and aneuploidy of chromosomes 18, X, and Y. Further details are available upon request. The second half of each trophectoderm biopsy (unused by FISH) from the first of 2 IVF cycles, as well as the arrested embryos from both cycles, and the entire trophectoderm biopsy of embryos from the second cycle were placed into polymerase chain reaction (PCR) tubes for cell lysis as previously described (16). Each lysate was split into 2 PCR tubes. One tube was processed by microarray analysis of [1] aneuploidy of all 24 chromosomes as previously described (16), [2] unbalanced derivative chromosomes as previously described (17), and [3] the microdeletion as described in Patient and Case Workup. The second PCR tube containing a split aliquot from the embryo lysates was processed by real-time PCR to determine the presence or absence of the microdeletion. Each SNP was genotyped as described in Patient and Case Workup. Newborn DNA DNA was isolated from a newborn buccal swab using the QIAamp DNeasy Tissue kit as recommended for cell cultures (QIAgen e60 Treff et al. Array-based balanced translocation test Vol. 96, No. 1, July 2011

4 Fertility and Sterility â e61 TABLE 2 Results of PGD for all embryos and tests used. Sample EM no. Microarray result FISH result TaqMan result rs rs rs rs rs rs Cycle 1 TE 1 45,XX, 13 Normal female CT TC GA TC AG AG TE 2 46,XY,t(2;20)(q21;p12.2) Male with ALGS T C A C G G TE 3 47,XX,þder(2)t(2;20)(q21;p12.2),þ17, 20 Unbalanced female T C A C G G TE 4 46,XX, 2,þder(20)t(2;20)(q21;p12.2) Indeterminate female CT TC GA TC AG AG TE 5 46,XX,t(2;20)(q21;p12.2) ALGS ND C A C G G TE 6 45,XX, 2 Unbalanced CT TC GA TC GA GA Arrested EM 7 46,XX,þ2, 20 T C A C G G Arrested EM 8 46,XX,der(20)t(2;20)(q21;p12.2) T C A C G G Arrested EM 9 46,XX,der(2)t(2;20)(q21;p12.2) CT CT AG CT AG AG Arrested EM 10 46,XY,der(20)t(2;20)(q21;p12.2) T ND A C G G Arrested EM 11 45,XY, 20 ND ND ND ND ND ND Arrested EM 12 46,XY,þder(2)t(2;20)(q21;p12.2), 20 T C A C G G Arrested EM 13 46,XY,þder(20)t(2;20)(q21;p12.2), 2 CT CT AG CT AG AG Arrested EM 14 45,XY,der(2)t(2;20)(q21;p12.2), 20 T C A C G G Arrested EM 15 46,XX,der(2)t(2;20)(q21;p12.2), 20 T C A C G G Arrested EM 16 47,XX,þ2,þ19, 20 T C A C G G Arrested EM 17 42,XY, 1, 5, 6,der(2)t(2;20)(q21;p12.2), T ND A ND G G þder(2)t(2;20)(q21;p12.2), 17, 20 Arrested EM 18 47,XY, 2,der(20)t(2;20)(q21;p12.2), CT CT AG CT AG AG þder(20)t(2;20)(q21;p12.2),þ20 Arrested EM 19 46,XY,der(2)t(2;20)(q21;p12.2), T C A C G G þder(2)t(2;20)(q21;p12.2), 12,þ16, 20 Cycle 2 TE 20 a 46,XY CT TC GA TC AG AG TE 21 46,XY (indeterminate q6) CT TC GA TC AG AG TE 22 45,XX,der(2)t(2;20)(q21;p12.2), 14 CT TC GA TC AG AG TE 23 46,XX, 2,þder(20)t(2;20)(q21;p12.2) CT TC GA TC AG AG TE 24 47,XY,þ15,der(20)t(2;20)(q21;p12.2) T C A C G G TE 25 45,XX,der(20)t(2;20)(q21;p12.2), 22 T C A C G G TE 26 46,XY,der(20)t(2;20)(q21;p12.2) T C A C G G TE 27 a 46,XY CT TC GA TC AG AG TE 28 46,XY, 2,der(20)t(2;20)(q21;p12.2), T C A C G G þder(20)t(2;20)(q21;p12.2) TE 29 45,XY, 20 T C A C G G TE 30 47,XY,der(20)t(2;20)(q21;p12.2),þ10 ND C A ND G G Arrested EM 31 47,XY, 2,þ16,þder(20)t(2;20)(q21;p12.2) CT TC GA TC AG AG Arrested EM 32 46,XY, 2,þder(20)t(2;20)(q21;p12.2) CT TC GA TC AG AG Arrested EM 33 45,XY, 6,der(20)t(2;20)(q21;p12.2) T C A C G G Arrested EM 34 46,XX,der(2)t(2;20)(q21;p12.2) CT TC GA TC AG AG Arrested EM 35 46XX,þder(2)t(2;20)(q21;12.2), 20 T C A C G G Arrested EM 36 47,XY,þ2,der(20)t(2;20)(q21;p12.2) ND CC AA CC GG GG Arrested EM 37 46,XX CT ND GA ND ND GG

5 TABLE 2 Continued. TaqMan result rs rs rs rs rs rs Sample EM no. Microarray result FISH result Arrested EM 38 46,XX, 1, 2,þ6,þder(20)t(2;20)(q21;p12.2) CT TC GA TC AG AG Arrested EM 39 46,XY,der(2)t(2;20)(q21;p12.2),þ20, 22 CT TC GA TC AG AG Arrested EM 40 46,XX,der(2)t(2;20)(q21;p12.2) CT TC GA TC AG AG Arrested EM 41 46,XX CT TC GA TC AG AG Arrested EM 42 No result CT ND GA CC ND GG Arrested EM 43 46,XY CT TC GA TC AG AG Arrested EM 44 45,XY,þder(2)t(2;20)(q21;p12.2), 16, 20 TT CC AA CC GG GG Arrested EM 45 46,XY, 1,þ10, 16,der(20)t(2;20)(q21;p12.2) TT CC AA CC GG GG Arrested EM 46 46,XY,der(20)t(2;20)(q21;p12.2) TT CC AA CC GG GG CT TC GA TC AG AG Arrested EM 47 47,XX, 2,der(20)t(2;20)(q21;p12.2), þder(20)t(2;20)(q21;p12.2),þ20 Arrested EM 48 46,XX, 4,þ8,t(2;20)(q21;p12.2) TT CC AA CC GG GG Note: EM ¼ embryo; TE ¼ trophectoderm. a Selected for transfer. Inc., Valencia, CA). Microarray detection of whole chromosome aneuploidy and microdeletions was conducted as described in Assays. ALGS real-time PCR was also performed as described in Assays. A conventional karyotype of the newborn was prepared by a cytogenetic reference laboratory. Ethics All materials were collected under institutional review board approval, and analysis was conducted with patient consent. RESULTS Microarray analysis revealed a 2.36-Mb microdeletion on the short arm of chromosome 20 in the patient with ALGS (Fig. 1B). Six real-time PCR based genotyping assays across the microdeletion were selected for further validation and were consistent with the original SNP microarray based genotyping (Table 1). Real-time PCR of preamplified DNA from maternal, paternal, and mixed maternal and paternal lymphocytes were also reliably and accurately genotyped (Table 1, Fig. 1C). In addition to the real-time PCR based detection of the microdeletion, a FISH probe for the Jagged1 locus was also validated by analysis of cells from the patient (Fig. 1D). The first treatment cycle resulted in the development of 19 embryos, of which 6 developed into morphologically normal cryopreservable blastocysts (Table 2). FISH analysis alone would have identified a single embryo as eligible for embryo transfer (Table 3). However, the embryo that was identified as eligible for transfer by FISH (embryo 1) displayed a monosomy of chromosome 13 by microarray (Fig. 2). With the criteria previously defined and validated for whole-chromosome aneuploidy (16), 2 embryos (embryo 2 and embryo 5) were diagnosed as either balanced or normal (Fig. 2). However, further analysis with the microdeletion settings (1-Mb smoothing) within the 20p12.2 segment resulted in these 2 embryos being diagnosed as abnormal (Fig. 2). Therefore, microarray analysis indicated that none of the embryos from the first treatment cycle were genetically normal. Although FISH was not as comprehensive as microarray, the 2 techniques were 100% consistent for the common regions that they assessed (Table 3). ALGS real-time PCR genotypes were consistent with expectations from microarray copy number assignments in all cases (Fig. 2). Overall, the ALGS real-time PCR based microdeletion predictions were 100% consistent with the SNP microarray and FISH-based microdeletion analyses (Table 3). Because FISH-based analysis was proven to be redundant in the presence of microarray- and real-time PCR based analyses (Table 3), embryos from the second treatment cycle were not screened by FISH. The second cycle led to the development of 29 embryos, of which 11 blastocysts were biopsied and cryopreserved (Table 2). Two of the 11 cryopreservable blastocysts were found to be euploid for all 24 chromosomes and negative for the presence of the microdeletion by both real-time PCR and microarray (Fig. 3A). Both embryos were thawed and transferred, resulting in the implantation and delivery of a genetically normal boy as confirmed by microarray (Fig. 3A), real-time PCR (Fig. 3B), and conventional karyotyping (Fig. 3C) of the newborn. DISCUSSION This is the first clinical case of microarray-based PGD being used to distinguish between balanced and normal chromosomes in embryos from a translocation carrier. Initial experience with SNP e62 Treff et al. Array-based balanced translocation test Vol. 96, No. 1, July 2011

6 TABLE 3 PGD results from cycle 1. Sample EM no. Microarray result FISH result TaqMan result rs rs rs rs rs rs TE 1 45,XX, 13 Normal female CT TC GA TC AG AG TE 2 46,XY,t(2;20)(q21;p12.2) Male with ALGS T C A C G G TE 3 47,XX,þder(2)t(2;20)(q21;p12.2), Unbalanced T C A C G G þ17, 20 female TE 4 46,XX, 2, Indeterminate CT TC GA TC AG AG þder(20)t(2;20)(q21;p12.2) female TE 5 46,XX,t(2;20)(q21;p12.2) ALGS ND C A C G G TE 6 45,XX, 2 Unbalanced CT TC GA TC GA GA Note: TE ¼ trophectoderm. microarray based aneuploidy and translocation screening indicated that 32% of developmentally competent blastocysts were balancednormal and euploid (17). Interestingly, analysis of DNA from the 17 patients who were translocation carriers did not reveal microdeletions near the breakpoints of either of the 2 chromosomes involved in the translocation. This was not unexpected given the lack of a clinical phenotype in those patients. The methodology described here may be more applicable to cases in which an apparently balanced FIGURE 2 (A) Microarray analysis of embryos derived from the first IVF cycle of the patient with a balanced translocation, 46,XX,t(2;20)(q21;p12.2), and ALGS caused by a microdeletion of 20p12.2. The 2 embryos (embryo 2 and embryo 5) diagnosed as either balanced or normal using the predefined chromosome aneuploidy analysis settings were subsequently identified as balanced ALGS carriers after the microdeletion analysis settings were used within 20p12. (B) Real-time PCR based analysis of the microdeletion was consistent with SNP microarray results. Fertility and Sterility â e63

7 FIGURE 3 SNP microarray and real-time PCR based analysis result (rs ) of (A) 2 embryos selected for transfer and (B) the resulting newborn. Both embryos and the resulting newborn were euploid and negative for the microdeletion. The newborn was found to be unaffected by ALGS and karyotypically normal by conventional G-banding analysis (C) and represents the first successful outcome following PGD to distinguish balanced from normal chromosomes in embryos from a translocation carrier. translocation has resulted in manifestation of a clinical disorder (i.e., ALGS). However, recently described methods of SNP microarray based haplotyping (19) might be used to distinguish between normal and balanced chromosomes in embryos from phenotypically normal translocation carriers. The data set described here represents an important and unique resource to validate such methodologies because it is the only SNP microarray data from embryos with known normal or balanced karyotypes. Therefore, the present study may represent an important step toward more generalized application of PGD to distinguish between balanced and normal chromosomes in embryos from translocation carriers. Although this advance may not be critical to the immediate goal of producing a healthy child for most translocation carriers, it may represent an important option for these patients to prevent their children from facing the same difficulties with reproduction in the future. Acknowledgment: The authors thank Lesley Northrop, Ph.D., for help with depositing the microarray data and preparation of the figures and tables. e64 Treff et al. Array-based balanced translocation test Vol. 96, No. 1, July 2011

8 REFERENCES 1. Van Dyke D, Weiss L, Roberson J, Babu V. The frequency and mutation rate of balanced autosomal rearrangements in man estimated from prenatal genetic studies for advanced maternal age. Am J Hum Genet 1983;35: Simovich MJ, Yatsenko SA, Kang SHL, Cheung SW, Dudek ME, Pursley A, et al. Prenatal diagnosis of a 9q34.3 microdeletion by array-cgh in a fetus with an apparently balanced translocation. Prenat Diagn 2007;27: Shanske AL, Edelmann L, Kardon NB, Gosset P, Levy B. Detection of an interstitial deletion of 2q21 22 by high resolution comparative genomic hybridization in a child with multiple congenital anomalies and an apparent balanced translocation. Am J Med Genet 2004;131A: Astbury C, Christ LA, Aughton DJ, Cassidy SB, Kumar A, Eichler EE, et al. Detection of deletions in de novo balance chromosome rearrangements: further evidence for their role in phenotypic abnormalities. Gene Med 2004;6: Kirchhoff M, Rose H, Lundsteen C. High resolution comparative genomic hybridization in clinical cytogenetics. J Med Genet 2001;38: Borg I, Squire M, Menzel C, Stout K, Morgan D, Willatt L, et al. A cryptic deletion of 2q35 including part of the PAX3 gene detected by breakpoint mapping in a child with autism and a de novo 2;8 translocation. J Med Genet 2002;39: Borg K, Stankiewicz P, Bocian E, Kruczek A, Obersztyn E, Lupski J, et al. Molecular analysis of a constitutional complex genome rearrangement with 11 breakpoints involving chromosomes 3, 11, 12, and 21 and a 0.5-Mb submicroscopic deletion in a patient with mild mental retardation. Hum Genet 2005;118: Vissers L, de Vries B, Osoegawa K, Janssen I, Feuth T, Choy C, et al. Array-based comparative genomic hybridization for the genomewide detection of submicroscopic chromosomal abnormalities. Am J Hum Genet 2003;73: Tyson C, McGillivray B, Chijiwa C, Rajcan- Separovic E. Elucidation of a cryptic interstitial 7q31.3 detection in a patient with a language disorder and mild mental retardation by array-cgh. Am J Hum Genet Part A 2004;129A: Shaw-Smith C, Redon R, Rickman L, Rio M, Willatt L, Fiegler H, et al. Microarray based comparative genomic hybridisation (array-cgh) detects submicroscopic chromosomal deletions and duplications in patients with learning disability/mental retardation and dysmorphic features. J Med Genet 2004;41: Gribble S, Prigmore E, Burford D, Porter K, Bee Ling N, Douglas E, et al. The complex nature of constitutional de novo apparently balanced translocations in patients presenting with abnormal phenotypes. J Med Genet 2005;42: Cheung S, Shaw C, Yu W, Li J, Ou Z, Patel A, et al. Development and validation of a CGH microarray for clinical cytogenetic diagnosis. Genet Med 2005;7: Krantz ID, Rand EB, Genin A, Hunt P, Jones M, Louis AA, et al. Deletions of 20p12 in Alagille syndrome: frequency and molecular characterization. Am J Med Genet 1997;70: Oda T, Elkahloun AG, Meltzer PS, Okajima K, Sugiyama K, Wada Y, et al. Identification of a larger than 3 Mb deletion including JAG1 in an Alagille syndrome patient with a translocation t(3;20)(q13.3;p12.2). Hum Mutat 2000; 16: Hattori M, Miyake Y, Oda T, Okajima K, Mizuno K, Sugiyama K, et al. Alagille syndrome with t(3;20)(q13.3;p12.2). Nippon Shonika Gakkai Zasshi 1995;99: Treff NR, Su J, Tao X, Levy B, Scott RT Jr. Accurate single cell 24 chromosome aneuploidy screening using whole genome amplification and single nucleotide polymorphism microarrays. Fertil Steril 2010;94: Treff NR, Northrop LE, Kasabwala K, Su J, Levy B, Scott RT Jr. Single nucleotide polymorphism microarray-based concurrent screening of 24 chromosome aneuploidy and unbalanced translocations in preimplantation human embryos. Fertil Steril 2011;95: e Treff NR, Su J, Kasabwala N, Miller KA, Levy B, Scott RT. Robust embryo identification using first polar body single nucleotide polymorphism (SNP) microarray-based DNA fingerprinting. Fertil Steril 2010;93: Handyside AH, Harton GL, Mariani B, Thornhill AR, Affara NA, Shaw MA, et al. Karyomapping: a universal method for genome wide analysis of genetic disease based on mapping crossovers between parental haplotypes. J Med Genet 2010;47: Fertility and Sterility â e65