Transcriptional sequencing: A method for DNA sequencing using RNA polymerase.
We have developed a sequencing method based on the RNA polymerase chain termination reaction with rhodamine dye attached to 3'-deoxynucleoside triphosphate (3'-dNTP). This method enables us to conduct a rapid isothermal sequencing reaction in <30 min, to reduce the amount of template required, and to do PCR direct sequencing without the elimination of primers and 2'-dNTP, which disturbs the Sanger sequencing reaction. An accurate and longer read length was made possible by newly designed four-color dye-3'-dNTPs and mutated RNA polymerase with an improved incorporation rate of 3'-dNTP. This method should be useful for large-scale sequencing in genome projects and clinical diagnosis.
Sasaki
N
N
Genome Science Laboratory, Tsukuba Life Science Center, The Institute of Physical and Chemical Research, 3-1-1 Koyadai, Tsukuba, Ibaraki 305, Japan.
Izawa
M
M
Watahiki
M
M
Ozawa
K
K
Tanaka
T
T
Yoneda
Y
Y
Matsuura
S
S
Carninci
P
P
Muramatsu
M
M
Okazaki
Y
Y
Hayashizaki
Y
Y
D004247:DNA / Q000235:genetics*; D004271:DNA, Fungal / Q000235:genetics; D012321:DNA-Directed RNA Polymerases / Q000235:genetics*; D016678:Genome; D012441:Saccharomyces cerevisiae / Q000235:genetics; D017422:Sequence Analysis, DNA / Q000379:methods*; D014158:Transcription, Genetic
D004271:DNA, Fungal; D004247:DNA; D012321:DNA-Directed RNA Polymerases