title: DNA template amplification genetic studies on archival human preimplantation biopsied microtubed cell samples.
abstract: Biopsied cell samples can remain in storage following an initial, unequivocally successful preimplantation genetic diagnosis (PGD). The fidelity of the DNA template for polymerase chain reaction (PCR) amplification of microtubed human preimplantation embryonic cells stored at -70 degrees C for extended periods of time (6 months to 4 years) has been assessed. PCR protocols and specific nested primer sets for the beta-globin, ZP3 and CA repeat motif successfully used for previous human embryo PGD research were employed for these studies. The results show that the DNA template of microtubed biopsied blastomere and mural trophectoderm cell samples stored for periods of up to 4 years may be successfully amplified by PCR. Specific gene sequences were able to be analysed at the 1-2 or 3-5 biopsied cell level with a 71-100% success rate. Analysis of DNA fragments amplified from the CA dinucleotide repeat locus showed that in 8/9 samples both alleles were amplified at the cellular level. No DNA contamination was detected in the stored microtubed samples, or in the experimental controls.
authors.affiliation: Division of Obstetrics and Gynaecology, UMDS, St. Thomas's Hospital, London, UK.
authors.forename: T A
authors.initials: TA
authors.lastname: Smith
authors.forename: M B
authors.initials: MB
authors.lastname: Wall
authors.forename: A L
authors.initials: AL
authors.lastname: Muggleton-Harris
mesh_terms: D000199:Actins / Q000235:genetics; D001706:Biopsy; D004247:DNA / Q000032:analysis / Q000235:genetics; D017931:DNA Primers; D004527:Egg Proteins / Q000235:genetics; D005260:Female; D005615:Freezing; D005914:Globins / Q000235:genetics; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D011247:Pregnancy; D019836:Preimplantation Diagnosis / Q000379:methods*; D011956:Receptors, Cell Surface; D013698:Templates, Genetic; D013997:Time Factors; D000072216:Zona Pellucida Glycoproteins
chemical_list: D000199:Actins; D017931:DNA Primers; D004527:Egg Proteins; D008562:Membrane Glycoproteins; D011956:Receptors, Cell Surface; C000606517:ZP3 protein, human; D000072216:Zona Pellucida Glycoproteins; D005914:Globins; D004247:DNA