Abstract
Contamination with extraneous DNA sequences is a frequent problem when performing PCR analysis of single cells. This report describes our experience with eliminating contaminating DNA sequences from PCR reagents for the purposes of gender identification. We have used amplification of Y-specific sequences to identify the gender of single human amniocytes. Female cells consistently showed no Y-specific bands but only 80% of male cells showed the expected intense Y-specific band. This phenomenon could lead to incorrect gender identification of single cells. We developed a technique of simultaneous amplification of X- and Y-specific sequences to prevent misdiagnosis because of failed PCR, which allows accurate preimplantation gender determination for women at risk for conceiving children with X-linked genetic discoses. We analyzed the gender of 141 consecutive single cells in a blinded manner without a single incorrect gender assignment
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Verlinsky Y, Pergament E, Binor Z, Rawlins R: Genetic analysis of human embryos prior to implantation: Future applications of in-vitro fertilization in treatment and prevention of human genetic diseases.In Future Aspects in Human in Vitro Fertilization, Ferchlinger, Kemeter (eds). Berlin, Springer-Verlag, pp 262–266
Penketh R, McLaren A: Prospects for prenatal diagnosis during preimplantation period. Bailliere's Clin Obstet Gynecol 1987;1(3):747–74
Coutell C, Williams C, Handyside A, Hardy K, Winston R, Williamson R: Genetic analysis of DNA from single human oocytes: A model for preimplantation diagnosis of cystic fibrosis. BMJ 1989;299:22–24
Handyside AH, Pattinson JK, Penketh RJA, Delhanty JDA, Winston RML, Tuddenham EGD: Biopsy of human preimplantation embryos and sexing by DNA and amplication. Lancet 1989;1:347–349
Handyside AH, Penketh RJA, Winston RML, Pattinson JK, Delhanty JDA, Tudenham EGD: Biopsy of human preimplantation embryos and sexing by DNA amplification. Lancer 1989;1(8634):347–349
Handyside AH, Kontogianni EH, Hardy K, Winston RML: Pregnancies from biopsied human preimplantation embryos sexed by Y-specific DNA amplification. Nature 1990;344:768–770
Strom CM, Verlinsky Y, Milayeva S, Evsikov S, Cieslak J, Lifchez A, Valle J, Moise J, Ginsberg N, Applebaum M: Preconception genetic diagnosis for cystic fibrosis by polar body removal and DNA analysis. Lancet 1990;336(8710):306–307
Verlinsky Y, Pergament E, Strom C: The preimplantation genetic diagnosis of genetic diseases. J Vitro Fert Embryo Transfer 1990;7:1–5
Verlinsky Y, Ginsberg N, Lifchez A, Valle J, Moise J, Strom CM: Analysis of the first polar body: Preconceptual genetic diagnosis. Hum Reprod 1990;7:826–829
Li HH, Gyllensten UB, Cui XF, Saikai RK, Ehrlich HA, Arnheim N: Amplification and analysis of DNA sequences in single human sperm and diploid cells. Nature 1988;335(6189):414–417
Saikai RK, Chang CA, Levensen CH, Warren TC, Boehm CD, Kazazian HH, Erlich HA: Diagnosis of sickle cell anemia and beta-thalassemia with enzymatically amplified DNA and nonradioactive allele-specific oligonucleotide probes. N Engl J Med 1988;319:537–541
Saikai RK, Scharf S, Faloona F, Mullis KB, Horn GT, Erlich HA, Arnheim N: Enzymatic amplification of beta-globin genomic sequences and restriction site analysis of diagnosis of sickle cell anemia. Science 1985;230:1350–1354
Chehab FF, Doherty M, Cai S, Kan YW, Cooper S, Rubin EM: Detection of sickle cell anaemia and thalessemias. Science 1987;329:293–294
Myerowitz R: Splice junction mutation in some Ashkenazi Jews with Tay-Sachs disease: Evidence against a single effect within this ethnic group. Proc Natl Acad Sci USA 1988;85(11):3955–3959
Myerowitz R, Costigan FC: The major defect in Ashkenazi Jews with Tay-Sachs disease is an insertion in the gene for the alpha-chain of beta-hexosaminidase. J Biol Chem 1988;00:000-000
Dermer SJ, Johnson EM: Rapid DNA analysis of alpha-1-antitrypsin deficiency: Application of an improved method for amplifying mutated gene sequences. Lab Invest 1988; 59:403–408
Speer A, Rosenthal A, Billwitz H, Hanke R: DNA amplification of a further exon of DuChenne muscular dystrophy locus increase possibilities for deletion screening. Nucleic Acids Res 1989;17(16):6774–6784
Kerem BE, Rommens JM, Buchanan JA, Markiewicz D, Cox TK, Chakravarti A, Buchwald M, Tsiu LC: Identification of the cystic fibrosis gene: Genetic analysis. Science 1989;245:1073–1080
Nakahori Y, Mitani K, Yamada M, Nakagome Y: A human Y-chromosome specific repeated DNA family (DYZ1) consists of a tandem array of pentanucleotides. Nucleic Acids Res 1986;14(19):7569–7579
Kogen SC, Doherty M, Gitschier J: An improved method for prenatal diagnosis of genetic diseases by analysis of amplified DNA sequences. N Engl J Med 317(16):985–990
Witt M, Erickson RP: A rapid method for detection of Y-chromosomal DNA from dried blood specimens by the polymerase chain reaction. Hum Genet 1989;82:271–274
Waye JS, Willard HF: Chromosome specific alpha satellite DNA: Nucleotide sequence analysis of the 2.0 kilobase-pair repeat from the human X chromosome. Nucleic Acids Res 1985;13(8):2731–2743
Porter-Jordan K, Garrett C: Source of contamination in polymerase chain reaction assay. Lancet 1990;May 19:1220
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Strom, C.M., Rechitsky, S. & Verlinsky, Y. Reliability of gender determination using the polymerase chain reaction (PCR) for single cells. J Assist Reprod Genet 8, 225–229 (1991). https://doi.org/10.1007/BF01130810
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF01130810