title: The comparison of the effectiveness of a modified conformation sensitive gel electrophoresis with denaturing high performance liquid chromatography.
abstract: Several methods have been developed for detection of sequence variation in genes and each has its advantages and disadvantages. A disadvantage of them is that the simpler, cost-effective methods are commonly perceived as being less sensitive in their detection of sequence variation, whereas those with proven sensitivity have a requirement for complex or expensive laboratory equipment. In this context, we undertook improvements to the conformation sensitive gel electrophoresis (CSGE) method which provides a cost-effective approach to mutation detection and compared the results with scanning carried out using denaturing high performance liquid chromatography (DHPLC) which utilises a dedicated analyser.
  
  
  
  We designed PCR primers to amplify the seven protein-coding exons of the human SPP2 gene which encodes secreted phosphoprotein 24 (spp24) such that the amplified products included the immediately-adjacent intronic regions. Five improvements were made to the CSGE method that was used to the scan the PCR-amplified DNA. The scanning was then repeated using DHPLC and the results were compared.
  
  
  
  Using CSGE, a single nucleotide polymorphism was discovered in exon 2 and another in intron 2 of the gene. Re-scanning of the same regions by DHPLC detected no additional sequence polymorphisms.
  
  
  
  With modification of the original protocol, CSGE is capable of providing a simple and cost-effective approach to the detection of DNA sequence polymorphisms that appears to be comparable in sensitivity to DHPLC.
authors.affiliation: Genetic Research Centre, the Social Welfare and Rehabilitation Sciences University, Tehran, Iran.|Dept.  of Genetics, University of Leicester, Leicester, UK.
authors.forename: Hamid Reza
authors.initials: HR
authors.lastname: Khorram Khorshid
authors.affiliation: Dept.  of Genetics, University of Leicester, Leicester, UK.
authors.forename: Raymond
authors.initials: R
authors.lastname: Dalgleish
mesh_terms: D001483:Base Sequence / Q000235:genetics; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004247:DNA / Q000737:chemistry* / Q000235:genetics*; D004252:DNA Mutational Analysis; D004587:Electrophoresis, Agar Gel / Q000379:methods; D005091:Exons / Q000235:genetics; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D010744:Phosphoric Monoester Hydrolases / Q000235:genetics; D017354:Point Mutation / Q000235:genetics
chemical_list: D008565:Membrane Proteins; D004247:DNA; C498526:SGPP2 protein, human; D010744:Phosphoric Monoester Hydrolases