The mutagenic properties of BrdUTP in a random mutagenesis process |
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Authors: | Xiangdong Ma Tao Ke Peihong Mao Xiang Jin Lixin Ma Guangyuan He |
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Institution: | (1) College of Life Science, Hubei University, Wuhan, 430062, China;(2) College of Life Science and Technology, Nan Yang Normal University, Nan Yang, 473061, China;(3) Ion Beam Biotechnology Center, Xinjiang University, Urumqi, Xinjiang, 830008, China;(4) China-UK HUST-RRes Crop Genetic Engineering and Genomics Joint Laboratory, College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, 430074, China |
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Abstract: | To explore the mutagenic properties of the nucleotide analogue bromodeoxyuridine triphosphate (BrdUTP), the wild type α-amylase
(xamy) gene from Xanthomonas campestris pv. campestris 8004 was used as a mutational target. It was mutated using PCR techniques to partially replace deoxythymidine triphosphate
(dTTP) with BrdUTP. A total of 18 mutants were selected for DNA sequencing from the mutagenesis libraries by their ability
to hydrolyze the starch. The results showed that 70% of the total mutations were single base-pair substitutions; BrdUTP also
induced deletion and insertion mutation types. Among single base-pair substitutions, the predominant mutation type is transition
(84%), but three kinds of transversions (16%) were also detected. It thus mainly induces A:T → G:C and T:A → C:G transitions.
This result indicated that when bromouracil is present as a deoxyribonucleoside triphosphate substrate it mainly paired with
dAMP, and when it is present as a template base it could pair with free dGTP. Three mutational hot spots induced by BrdUTP
were revealed in this work. |
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Keywords: | BrdUTP Mutagenic properties PCR Taq DNA polymerase |
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