scholarly journals Real-Time Monitoring of DNA Polymerase Function and Stepwise Single-Nucleotide DNA Strand Translocation through a Protein Nanopore

2010 ◽  
Vol 49 (52) ◽  
pp. 10106-10109 ◽  
Author(s):  
John Chu ◽  
Marcos González-López ◽  
Scott L. Cockroft ◽  
Manuel Amorin ◽  
M. Reza Ghadiri
2010 ◽  
Vol 122 (52) ◽  
pp. 10304-10307 ◽  
Author(s):  
John Chu ◽  
Marcos González-López ◽  
Scott L. Cockroft ◽  
Manuel Amorin ◽  
M. Reza Ghadiri

2006 ◽  
Vol 353 (1) ◽  
pp. 141-143 ◽  
Author(s):  
Changbei Ma ◽  
Zhiwen Tang ◽  
Kemin Wang ◽  
Weihong Tan ◽  
Jun Li ◽  
...  

2016 ◽  
Vol 113 (19) ◽  
pp. 5233-5238 ◽  
Author(s):  
Carl W. Fuller ◽  
Shiv Kumar ◽  
Mintu Porel ◽  
Minchen Chien ◽  
Arek Bibillo ◽  
...  

DNA sequencing by synthesis (SBS) offers a robust platform to decipher nucleic acid sequences. Recently, we reported a single-molecule nanopore-based SBS strategy that accurately distinguishes four bases by electronically detecting and differentiating four different polymer tags attached to the 5′-phosphate of the nucleotides during their incorporation into a growing DNA strand catalyzed by DNA polymerase. Further developing this approach, we report here the use of nucleotides tagged at the terminal phosphate with oligonucleotide-based polymers to perform nanopore SBS on an α-hemolysin nanopore array platform. We designed and synthesized several polymer-tagged nucleotides using tags that produce different electrical current blockade levels and verified they are active substrates for DNA polymerase. A highly processive DNA polymerase was conjugated to the nanopore, and the conjugates were complexed with primer/template DNA and inserted into lipid bilayers over individually addressable electrodes of the nanopore chip. When an incoming complementary-tagged nucleotide forms a tight ternary complex with the primer/template and polymerase, the tag enters the pore, and the current blockade level is measured. The levels displayed by the four nucleotides tagged with four different polymers captured in the nanopore in such ternary complexes were clearly distinguishable and sequence-specific, enabling continuous sequence determination during the polymerase reaction. Thus, real-time single-molecule electronic DNA sequencing data with single-base resolution were obtained. The use of these polymer-tagged nucleotides, combined with polymerase tethering to nanopores and multiplexed nanopore sensors, should lead to new high-throughput sequencing methods.


2019 ◽  
Vol 142 ◽  
pp. 111528 ◽  
Author(s):  
Johanna Roether ◽  
Kang-Yu Chu ◽  
Norbert Willenbacher ◽  
Amy Q. Shen ◽  
Nikhil Bhalla

2016 ◽  
Vol 408 (12) ◽  
pp. 3275-3280 ◽  
Author(s):  
Changbei Ma ◽  
Haisheng Liu ◽  
Jun Wang ◽  
Shunxin Jin ◽  
Kemin Wang

1992 ◽  
Vol 64 (17) ◽  
pp. 1996-1997 ◽  
Author(s):  
Toshinari. Sakurai ◽  
Yuzuru. Husimi

2006 ◽  
Vol 175 (4S) ◽  
pp. 521-521
Author(s):  
Motoaki Saito ◽  
Tomoharu Kono ◽  
Yukako Kinoshita ◽  
Itaru Satoh ◽  
Keisuke Satoh

2001 ◽  
Vol 11 (PR3) ◽  
pp. Pr3-1175-Pr3-1182 ◽  
Author(s):  
M. Losurdo ◽  
A. Grimaldi ◽  
M. Giangregorio ◽  
P. Capezzuto ◽  
G. Bruno

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