PMID- 10220146
OWN - NLM
STAT- MEDLINE
DCOM- 19990610
LR  - 20061115
IS  - 1059-7794 (Print)
IS  - 1059-7794 (Linking)
VI  - 13
IP  - 4
DP  - 1999
TI  - High-throughput single-strand conformation polymorphism analysis by automated
      capillary electrophoresis: robust multiplex analysis and pattern-based
      identification of allelic variants.
PG  - 318-27
AB  - Genetic diagnosis of an inherited disease or cancer often involves analysis for
      unknown point mutations in several genes; therefore, rapid and automated
      techniques that can process a large number of samples are needed. We describe a
      method for high-throughput single-strand conformation polymorphism (SSCP)
      analysis using automated capillary electrophoresis. The operating temperature of 
      a commercially available capillary electrophoresis instrument (ABI PRISM 310) was
      expanded by installation of a cheap in-house designed cooling system, thereby
      allowing us to perform automated SSCP analysis at 14-45 degrees C. We have used
      the method for detection of point mutations associated with the inherited cardiac
      disorders long QT syndrome (LQTS) and hypertrophic cardiomyopathy (HCM). The
      sensitivity of the method was 100% when 34 different point mutations were
      analyzed, including two previously unpublished LQTS-associated mutations (F157C
      in KVLQT1 and G572R in HERG), as well as eight novel normal variants in HERG and 
      MYH7. The analyzed polymerase chain reaction (PCR) fragments ranged in size from 
      166 to 1,223 bp. Seventeen different sequence contexts were analyzed. Three
      different electrophoresis temperatures were used to obtain 100% sensitivity. Two 
      mutants could not be detected at temperatures greater than 20 degrees C. The
      method has a high resolution and good reproducibility and is very robust, making 
      multiplex SSCP analysis and pattern-based identification of known allelic
      variants as single nucleotide polymorphisms (SNPs) possible. These possibilities,
      combined with automation and short analysis time, make the method suitable for
      high-throughput tasks, such as genetic screening.
FAU - Larsen, L A
AU  - Larsen LA
AD  - Department of Clinical Biochemistry, Statens Serum Institut, Copenhagen, Denmark.
FAU - Christiansen, M
AU  - Christiansen M
FAU - Vuust, J
AU  - Vuust J
FAU - Andersen, P S
AU  - Andersen PS
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - United States
TA  - Hum Mutat
JT  - Human mutation
JID - 9215429
SB  - IM
MH  - *Alleles
MH  - Cardiomyopathy, Hypertrophic/genetics
MH  - DNA Mutational Analysis/instrumentation/*methods
MH  - Electrophoresis, Capillary/instrumentation/*methods
MH  - Humans
MH  - Long QT Syndrome/genetics
MH  - Point Mutation
MH  - Polymerase Chain Reaction
MH  - *Polymorphism, Genetic
MH  - *Polymorphism, Single-Stranded Conformational
MH  - Sequence Analysis, DNA
MH  - Temperature
EDAT- 1999/04/29 02:03
MHDA- 2000/06/22 10:00
CRDT- 1999/04/29 02:03
PHST- 1999/04/29 02:03 [pubmed]
PHST- 2000/06/22 10:00 [medline]
PHST- 1999/04/29 02:03 [entrez]
AID - 10.1002/(SICI)1098-1004(1999)13:4<318::AID-HUMU9>3.0.CO;2-F [pii]
AID - 10.1002/(SICI)1098-1004(1999)13:4<318::AID-HUMU9>3.0.CO;2-F [doi]
PST - ppublish
SO  - Hum Mutat. 1999;13(4):318-27. doi:
      10.1002/(SICI)1098-1004(1999)13:4<318::AID-HUMU9>3.0.CO;2-F.