Open AccessTechnical report: Part 2. Basic requirements for designing optimal PCR primers
Author(s) -
Mitsuhashi Masato
Publication year - 1996
Publication title -
journal of clinical laboratory analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.536
H-Index - 50
eISSN - 1098-2825
pISSN - 0887-8013
DOI - 10.1002/(sici)1098-2825(1996)10:5<285::aid-jcla9>3.0.co;2-7
Subject(s) - in silico pcr , primer (cosmetics) , amplicon , multiplex polymerase chain reaction , primer dimer , polymerase chain reaction , biology , microbiology and biotechnology , applications of pcr , multiplex , polymerase chain reaction optimization , cloning (programming) , computational biology , nested polymerase chain reaction , inverse polymerase chain reaction , genetics , gene , chemistry , computer science , organic chemistry , programming language
Designing optimal polymerase chain reaction (PCR) primer sequences is one of the critical factors for successful PCR with sensitive, specific, and assay‐to‐assay reproducible results. In this review, all the requirements of PCR primer sequences are summarized, such as location, size of amplicon, length of primers, nucleotide composition, Tm, 3′ terminal hybridization strength and frequency, hairpin formation energy, primer‐to‐primer interaction, specificity, and location of mismatches to sequences of cross‐hybridization. The report also discusses how to explore these various types of information for more advanced PCR applications, which include nested PCR, multiplex PCR, competitive PCR, long PCR, point mutation detection, degenerate primers, and PCR cloning. © 1996 Wiley‐Liss, Inc.