Recycling of a single human blastomere fixed on a microscopic slide for sexing and diagnosis of specific mutations by various types of polymerase chain reaction.
OBJECTIVE
To investigate the suitability of recycling single blastomeres to assess multiple genetic variables for preimplantation genetic diagnosis.


METHODS
Prospective randomized study.


METHODS
An academic medical center.


METHODS
Patients undergoing IVF-ET.


METHODS
Blastomeres were disaggregated from donated embryos obtained from patients.


METHODS
Polymerase chain reaction (PCR) amplification products.


RESULTS
Fifty-eight blastomeres individually fixed on slides were separated into four groups. Sequential PCRs (group I, n = 30), primed in situ labeling (PRINS) before five sequential PCRs (group II, n = 10), staining with hematoxylin before performing five sequential PCRs (group III, n = 11) and preamplification of whole DNAs by degenerate oligonucleotide primer (DOP) before performing PCR were executed. The amplification efficiencies of five sequential PCRs were 100%, 100%, 96.6%, 83.3%, 56.7% for group I; 100% 100%, 100%, 80%, 40% for group II; 54.5%, 36.4%, 18.2%, 9.1% for group III; and 100%, 100%, 100%, 100%, 100% for group IV.


CONCLUSIONS
Blastomeres fixed for PRINS can be recycled for PCR to obtain more genetic information. Hematoxylin staining appears to increase the incidence of failed amplification. Preamplification of whole genomic DNAs by DOP-PCR appears to facilitate diagnosis with high efficiency.
He
Z Y
ZY
Center for Reproductive Medicine and Infertility, Weill Medical College of Cornell University, New York, New York, USA.
Liu
H C
HC
Mele
C A
CA
Veeck
L L
LL
Davis
O
O
Rosenwaks
Z
Z
D001757:Blastomeres / Q000166:cytology* / Q000502:physiology*; D017931:DNA Primers; D004276:DNA, Satellite / Q000235:genetics; D004624:Embryo Transfer; D005260:Female; D005307:Fertilization in Vitro; D006801:Humans; D066298:In Vitro Techniques; D009154:Mutation; D016133:Polymerase Chain Reaction / Q000379:methods*; D011446:Prospective Studies; D012732:Sex Determination Analysis / Q000379:methods*; D014960:X Chromosome; D014998:Y Chromosome
D017931:DNA Primers; D004276:DNA, Satellite