Betaine, Dimethyl Sulfoxide, and 7-Deaza-dGTP, a Powerful Mixture for Amplification of GC-Rich DNA Sequences

• Published in: The Journal of molecular diagnostics : JMD Vol. 8; no. 5; pp. 544 - 550
• Main Authors: Musso, Marco, Bocciardi, Renata, Parodi, Sara, Ravazzolo, Roberto, Ceccherini, Isabella
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.11.2006; ASIP; American Society for Investigative Pathology
• Subjects: Base Sequence; Betaine - chemistry; Deoxyguanine Nucleotides - chemistry; Dimethyl Sulfoxide - chemistry; DNA - metabolism; GC Rich Sequence; Gene Amplification; Homeodomain Proteins - genetics; Humans; LIM-Homeodomain Proteins; Molecular Sequence Data; Polymerase Chain Reaction - methods; Polymerase Chain Reaction - standards; Promoter Regions, Genetic; Proto-Oncogene Proteins c-ret - genetics; Technical Advance; Transcription Factors - genetics
• ISSN: 1525-1578; 1943-7811
• DOI: 10.2353/jmoldx.2006.060058

Currently, polymerase chain reaction is the most used technique in many laboratories for either diagnostic or molecular biology purposes. Despite the large number of DNA sequences that can be easily analyzed, some GC-rich sequences are refractory to amplification due to the formation of secondary intramolecular structures. To overcome this problem, several molecules have been described to improve polymerization. Here we show that a combination of three additives—betaine, dimethyl sulfoxide, and 7-deaza-dGTP—was essential to achieve amplification of DNA sequences of three disease genes showing a GC content ranging from 67 to 79%.