Diagnosis of haploidy and triploidy based on measurement of gene copy number by real-time PCR

Hum Mutat. 2000 Nov;16(5):431-6. doi: 10.1002/1098-1004(200011)16:5<431::AID-HUMU8>3.0.CO;2-Z.


We report the development of a method for diagnosis of heterozygous deletions or duplications based on measurement of gene copy number. The method involves amplifications of a test locus with unknown copy number and a reference locus with known copy number using real-time PCR. Progress of the PCR reactions is monitored using fluorigenic probes and a real-time fluorescence detection system. For each reaction, the number of cycles is measured at which a defined threshold fluorescence emission is reached. Using standard curves, the copy number of the test DNA relative to a common standard DNA is determined for each locus. From the ratio of the relative copy numbers, the genomic copy number of the test locus is determined. In order to demonstrate the accuracy and reliability of the method for genetic testing, we analyzed 43 patients with hereditary neuropathy with liability to pressure palsies (HNPP), containing a heterozygous deletion of a 1.5 Mb region on chromosome 17p11.2-p12, eight patients with Charcot-Marie-Tooth disease, containing a heterozygous duplication of the same genomic region, and 50 normal control individuals. As a test locus we analyzed the PMP22 gene located within the 1.5 Mb region. The genomic copy number of the test locus was precisely measured, and the presence or absence of the genomic deletion or duplication was unambiguously diagnosed in all individuals.

Publication types

  • Comparative Study

MeSH terms

  • Charcot-Marie-Tooth Disease / diagnosis
  • Charcot-Marie-Tooth Disease / genetics
  • Computer Systems
  • Female
  • Gene Dosage*
  • Gene Duplication
  • Haploidy*
  • Hereditary Sensory and Motor Neuropathy / diagnosis
  • Hereditary Sensory and Motor Neuropathy / genetics
  • Heterozygote
  • Humans
  • Male
  • Myelin Proteins / genetics
  • Polymerase Chain Reaction* / instrumentation
  • Polymerase Chain Reaction* / methods
  • Polyploidy*
  • Sequence Deletion / genetics


  • Myelin Proteins
  • PMP22 protein, human