Fluorescent-based single-strand conformation polymorphism/heteroduplex capillary electrophoretic mutation analysis of the P53 gene.
Fluorescent-based single-strand conformation polymorphism (F-SSCP) analysis with capillary electrophoresis (CE) is the most common method for the detection of mutation because of its high sensitivity and resolution. In this study, we prepared an inexpensive linear polyacrylamide (LPA), and successfully applied it to CE-SSCP analysis and tandem CE-SSCP/heteroduplex analysis (HA) of the P53 gene on an ABI capillary genetic analyzer. A comparison of the sieving capabilities of a homemade LPA and commercial polydimethylacrylamide (PDMA) demonstrates that the homemade LPA has a higher resolution, a shorter analysis time, and is more suitable for tandem SSCP/HA than commercial PDMA. To show the usefulness, mutations of P53 gene exon 7 - 8 in 37 tumor samples were investigated by using homemade LPA. The results indicate that 10 mutations were found in 9 of 37 cases; the majority of P53 mutations were missense mutations, and 70% were located in exon 7, which plays an important role in neoplastic progression in human tumorigenesis.
National Chromatographic R&A Center, Dalian Institute of Chemical Physics, The Chinese Academy of Sciences, Dalian, China.
Chunxia
C
Zhao
Guowang
G
Xu
Xianzhe
X
Shi
Jianmei
J
Ma
Yan
Y
Zhang
Shen
S
Lv
Qianfeng
Q
Weng
Qing
Q
Yang
D000180:Acrylic Resins / Q000737:chemistry; D003062:Codon; D004252:DNA Mutational Analysis; D005091:Exons; D005456:Fluorescent Dyes / Q000494:pharmacology*; D016158:Genes, p53; D006801:Humans; D009154:Mutation; D020125:Mutation, Missense; D009692:Nucleic Acid Heteroduplexes / Q000737:chemistry; D016133:Polymerase Chain Reaction; D011108:Polymers / Q000737:chemistry; D018807:Polymorphism, Single-Stranded Conformational; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D013050:Spectrometry, Fluorescence / Q000379:methods*; D013997:Time Factors; D000099166:Polyacrylamides
D000180:Acrylic Resins; D003062:Codon; D005456:Fluorescent Dyes; D009692:Nucleic Acid Heteroduplexes; D011108:Polymers; D000099166:Polyacrylamides