A technique for determination of the base sequence of a homogeneous polynucleotide that acts as a template in a replication system. A small amount of a specific dideoxynucleoside triphosphate is included with the four normal deoxynucleotide triphosphates during replication. Chain termination occurs whenever a dideoxynucleotide is incorporated that generates a labelled polynucleotide whose chain length, evaluated by polyacrylamide-gel electrophoresis, is indicative of the distance from the 5'-end of the primer to the position at which the normal nucleotide occurs. Detection of the electrophoresis bands may be by a label such as 32P or, more recently, by a fluorophore incorporated into the primer (dye primer sequencing) or the dideoxynucleotide (dye terminator sequencing). In fact, in a variation of the dye terminator method, the labelling of each of the four dideoxynucleotides with a fluorophore that emits at a different wavelength permits the identification of the base that terminates each electrophoresis band within a separated mixture of all four (single lane sequencing). (see also Maxam-Gilbert method (chemical cleavage method); plus and minus method)Sanger, F., Nicklen, S. and Coulson, A.R. (1977) Proc. Natl. Acad. Sci. U.S.A. 74, 5463-5467
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GenSmart™ Design is a free online DNA construct design tool developed by GenScript. GenSmart™ Design has two design modules, the Create Construct module for individual plasmid design and the Create Library module for DNA library design.
This online tool shows commonly used genetic codon frequency table in expression host organisms including Escherichia coli and other common host organisms.
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