Detection of sequence variability of the collagen type IIalpha 1 3' variable number of tandem repeat.
The variable number of tandem repeat (VNTR) 3' of the collagen type II (COL2A1) gene has been shown to be highly variable with a complex molecular structure. In a previous pilot experiment we observed discordance between methods to genotype this informative marker. To further investigate the extent and molecular nature of this discordance, we genotyped a random sample of 207 Caucasian individuals with two genotyping methods and sequenced new alleles. We compared single-strand (SS) analysis, which is based on detection of size differences between the different alleles, and heteroduplex analysis (HA), which is sensitive to both size and sequence differences. Overall, 26% of discordance between the two methods was detected. Approximately two thirds of this discordance was caused by subdivision of SS-alleles 13R1 and 14R2 into HA-alleles 4A + 4B and 3B + 3C, respectively. Sequence analysis of the COL2A1 VNTR alleles 4B and 3C showed that these alleles differed in sequence, but not in size, from already described SS-alleles, which explains why they escape detection by SS. The 4B allele is a frequent allele in the population (14%) and is, therefore, important to distinguish in association studies. We conclude that HA is a reliable method when the described optimized electrophoretic conditions are used. HA is a sensitive genotyping method to document allelic diversity at this locus, which can distinguish more alleles compared to the SS method.
Department of Internal Medicine, Erasmus University Rotterdam, The Netherlands. vanmeurs@endov.fgg.eur.nl
J B
JB
van Meurs
P P
PP
Arp
Y
Y
Fang
P E
PE
Slagboom
I
I
Meulenbelt
J P
JP
van Leeuwen
H A
HA
Pols
A G
AG
Uitterlinden
D000368:Aged; D000483:Alleles; D001483:Base Sequence; D003094:Collagen / Q000235:genetics*; D004277:DNA, Single-Stranded / Q000032:analysis; D005260:Female; D014644:Genetic Variation; D020180:Heteroduplex Analysis; D006801:Humans; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D009692:Nucleic Acid Heteroduplexes / Q000032:analysis; D017422:Sequence Analysis, DNA; D020080:Tandem Repeat Sequences
D004277:DNA, Single-Stranded; D009692:Nucleic Acid Heteroduplexes; D003094:Collagen