Simple detection of large InDeLS by DHPLC: the ACE gene as a model.
Insertion-deletion polymorphism (InDeL) is the second most frequent type of genetic variation in the human genome. For the detection of large InDeLs, researchers usually resort to either PCR gel analysis or RFLP, but these are time consuming and dependent on human interpretation. Therefore, a more efficient method for genotyping this kind of genetic variation is needed. In this report, we describe a method that can detect large InDeLs by DHPLC (denaturating high-performance liquid chromatography) using the angiotensin-converting enzyme (ACE) gene I/D polymorphism as a model. The InDeL targeted in this study is characterized by a 288 bp Alu element insertion (I). We used DHPLC at nondenaturating conditions to analyze the PCR product with a flow through the chromatographic column under two different gradients based on the differences between D and I sequences. The analysis described is quick and easy, making this technique a suitable and efficient means for DHPLC users to screen InDeLs in genetic epidemiological studies.
Koyama
Renata Guedes
RG
Departamento de Psicobiologia, Universidade Federal de São Paulo (UNIFESP), Rua Napoleão de Barros, 925/1 andar Vila Clementino, São Paulo 04024-002, SP, Brazil.
Castro
Rosa M R P S
RM
De Mello
Marco Túlio
MT
Tufik
Sergio
S
Pedrazzoli
Mario
M
D000483:Alleles; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D005787:Gene Frequency; D015894:Genome, Human; D005838:Genotype; D006801:Humans; D016254:Mutagenesis, Insertional / Q000706:statistics & numerical data; D007703:Peptidyl-Dipeptidase A / Q000235:genetics*; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic; D012680:Sensitivity and Specificity; D016415:Sequence Alignment; D017384:Sequence Deletion
D007703:Peptidyl-Dipeptidase A