title: Oligonucleotide properties determination and primer designing: a critical examination of predictions.
abstract: BACKGROUND
  Precise prediction of melting temperature (T(m)), secondary structures and design of oligonucleotides determine the efficiency and success of experimentation in molecular biology. Availability of a plethora of software and the users unawareness about their limitations compromises the accuracy and reliability of the predictions.
  
  
  RESULTS
  Comparative analysis of 56 modules was done for T(m) prediction using a large set of oligonucleotide sequences spanning the whole range of GC-content and length. Allawi module of the calculator 'MELTING', Nearest Neighbor (NN) of oligo calculator (McLab), NN of T(m) Calculation for Oligos (Biomath Calculator, Promega) and HYTHER provided the most precise T(m) predictions. A model has also been proposed to calculate the optimum annealing temperature integrating the already reported formulations. Secondary structure predictions of oligonucleotides reveal a large number of structures in contrast to the experimental observations. Of the 11 primer designing tools evaluated, Primer 3 and WebPrimer performed the best for the AT-rich templates, Exon Primer for AT = GC templates, and Primer Design Assistant, Primer3 and Primer Quest for GC-rich templates. This study provides optimal choice for application to the user, increasing the success of a variety of experimentations, especially those that have high-throughput and complex assay designs.
  
  
  BACKGROUND
  db@igib.res.in
  
  
  BACKGROUND
  The details of the oligonucleotides and of the different modules of T(m) prediction considered for the study are provided as Supplementary Information, available at Bioinformatics online.
authors.affiliation: Functional Genomics Unit, Institute of Genomics and Integrative Biology, Delhi, India.
authors.forename: Sreenivas
authors.initials: S
authors.lastname: Chavali
authors.forename: Anubha
authors.initials: A
authors.lastname: Mahajan
authors.forename: Rubina
authors.initials: R
authors.lastname: Tabassum
authors.forename: Souvik
authors.initials: S
authors.lastname: Maiti
authors.forename: Dwaipayan
authors.initials: D
authors.lastname: Bharadwaj
mesh_terms: D003198:Computer Simulation; D017931:DNA Primers / Q000032:analysis* / Q000737:chemistry*; D008956:Models, Chemical; D008957:Models, Genetic; D016133:Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D012984:Software; D012986:Software Validation; D044366:Transition Temperature
chemical_list: D017931:DNA Primers