Differences in mitochondrial and nuclear DNA status of high-density and low-density sperm fractions after density centrifugation preparation.
OBJECTIVE
To determine the mitochondrial DNA and nuclear DNA fragmentation of sperm populations separated by using discontinuous density gradient.


METHODS
Analysis of mitochondrial and nuclear DNA status of sperm from high and low density layers.


METHODS
Regional fertility center.


METHODS
Twenty-eight men who presented for an initial infertility investigation.


METHODS
Semen was prepared by using discontinuous density gradient (90.0%:45.0%) and subjected to a modified long polymerase chain reaction to assess mitochondrial DNA deletions and to a modified single-cell alkaline gel electrophoresis assay to determine nuclear DNA fragmentation.


RESULTS
The high-density fraction displayed significantly more wild-type mitochondrial DNA (75% of samples) than did the low-density fraction (25% of samples). In the high-density fraction, the incidence of single deletions was higher than that of double or multiple deletions, and the deletions were predominantly small. A strong correlation was observed between nuclear DNA fragmentation and the number and size of mitochondrial DNA deletions.


CONCLUSIONS
Density centrifugation isolates a population of sperm with high-quality mitochondrial DNA and nuclear DNA.
O'Connell
Michael
M
Obstetrics and Gynaecology, Queen's University Belfast, Institute of Clinical Science and Royal Maternity Hospital, Belfast, United Kingdom
McClure
Neil
N
Powell
Lesley A
LA
Steele
E Kristine
EK
Lewis
Sheena E M
SE
D000328:Adult; D002499:Centrifugation, Density Gradient; D020552:Comet Assay; D053938:DNA Fragmentation / Q000502:physiology*; D004272:DNA, Mitochondrial / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D007248:Infertility, Male / Q000235:genetics* / Q000473:pathology; D008297:Male; D016133:Polymerase Chain Reaction; D013094:Spermatozoa / Q000002:abnormalities / Q000737:chemistry / Q000502:physiology*
D004272:DNA, Mitochondrial