The objective of this study was to identify twinning rate quantitative traitloci (QTL) by typing pooled samples in a preliminary screening followed byinterval mapping to test QTL effects. Four elite North American Holsteinhalf-sib sire families with high twinning rate predicted transmitting abilities(PTA) were used in this study. Chromosomes 5, 7, 19 and 23 were not genotyped asthese chromosomes were scanned for QTL in these families in a previous study.DNA was extracted from phenotypically extreme sons in each sire family. Twopools were prepared from sons of sires in each phenotypic tail, two each forhigh and low PTA levels for twinning rates. Each pool contained DNA from 4 to15% of all sons of the sire depending on family. A total of 268 fluorescentlylabelled microsatellite markers were tested for heterozygosity in sires. About135--170 informative markers per family were genotyped using pooled DNA samples.Based on the preliminary evidence for potential twinning rate QTL from pooledtyping, interval mapping was performed subsequently on 12 chromosomal regions byfamily combinations. Evidence of QTL for twinning rate was found in one familyon BTA 21 and 29 at a chromosome-wide P<0.05 and on BTA 8, 10 and 14 with achromosome-wide P<0.01.