title: Direct visualization of cystic fibrosis transmembrane regulator mutations in the clinical laboratory setting.
abstract: BACKGROUND
  The recommendation for population- based cystic fibrosis (CF) carrier screening by the American College of Medical Genetics for the 25 most prevalent mutations and 6 polymorphisms in the CF transmembrane regulatory gene has greatly increased clinical laboratory test volumes. We describe the development and technical validation of a DNA chip in a 96-well format to allow for high-throughput genotype analysis.
  
  
  METHODS
  The CF Portrait chip contains an 8 x 8 array of capture probes and controls to detect all requisite alleles. Single-tube multiplex PCR with 15 biotin-labeled primer pairs was used to amplify sequences containing all single-nucleotide polymorphisms to be interrogated. Detection of a thin-film signal created by hybridization of multiplex PCR-amplified DNA to complementary capture probes was performed with an automated image analysis instrument, NucleoSight. Allele classification, data formatting, and uploading to a laboratory information system were fully automated.
  
  
  RESULTS
  The described platform correctly classified all mutations and polymorphisms and can screen approximately 1300 patient samples in a 10-h shift. Final validation was performed by two separate 1000-sample comparisons with Roche CF Gold line probe strips and the Applera CF OLA, Ver 3.0. The CF Portrait Biochip made no errors during this validation, whereas the Applera assay made seven miscalls of the IVS-8 5T/7T/9T polymorphism
  
  
  CONCLUSIONS
  The CF Portrait platform is an automated, high-throughput, DNA chip-based assay capable of accurately classifying all CF mutations in the recommended screening panel, including the IVS-8 5T/7T/9T polymorphism.
authors.affiliation: Nichols Institute, Quest Diagnostics, San Juan Capistrano, CA, USA.
authors.forename: Charles M
authors.initials: CM
authors.lastname: Strom
authors.forename: David D
authors.initials: DD
authors.lastname: Clark
authors.forename: Feras M
authors.initials: FM
authors.lastname: Hantash
authors.forename: Larry
authors.initials: L
authors.lastname: Rea
authors.forename: Ben
authors.initials: B
authors.lastname: Anderson
authors.forename: Diana
authors.initials: D
authors.lastname: Maul
authors.forename: Donghui
authors.initials: D
authors.lastname: Huang
authors.forename: Donald
authors.initials: D
authors.lastname: Traul
authors.forename: Christina
authors.initials: C
authors.lastname: Chen Tubman
authors.forename: Renee
authors.initials: R
authors.lastname: Garcia
authors.forename: P Patrick
authors.initials: PP
authors.lastname: Hess
authors.forename: Hong
authors.initials: H
authors.lastname: Wang
authors.forename: Beryl
authors.initials: B
authors.lastname: Crossley
authors.forename: Evelyn
authors.initials: E
authors.lastname: Woodruff
authors.forename: Rebecca
authors.initials: R
authors.lastname: Chen
authors.forename: Myra
authors.initials: M
authors.lastname: Killeen
authors.forename: Weimin
authors.initials: W
authors.lastname: Sun
authors.forename: Jonathan
authors.initials: J
authors.lastname: Beer
authors.forename: Heather
authors.initials: H
authors.lastname: Avens
authors.forename: Barry
authors.initials: B
authors.lastname: Polisky
authors.forename: Robert D
authors.initials: RD
authors.lastname: Jenison
mesh_terms: D001322:Autoanalysis; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics*; D015342:DNA Probes; D005820:Genetic Testing / Q000379:methods; D005838:Genotype; D006801:Humans; D009154:Mutation; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D020641:Polymorphism, Single Nucleotide; D012371:Robotics
chemical_list: C505032:CFTR protein, human; D015342:DNA Probes; D019005:Cystic Fibrosis Transmembrane Conductance Regulator