scholarly journals Optimizing T4 DNA polymerase conditions enhances the efficiency of one-step sequence- and ligation-independent cloning

BioTechniques ◽  
2017 ◽  
Vol 63 (3) ◽  
Author(s):  
Mohammad Nazrul Islam ◽  
Kyeong Won Lee ◽  
Hyung-Soon Yim ◽  
Seong Hyuk Lee ◽  
Hae Chang Jung ◽  
...  
Keyword(s):  
Dna Polymerase ◽  
Ligation Independent Cloning ◽  
One Step

scholarly journals One-Step Sequence- and Ligation-Independent Cloning as a Rapid and Versatile Cloning Method for Functional Genomics Studies

2012 ◽  
Vol 78 (15) ◽  
pp. 5440-5443 ◽  
Author(s):  
Jae-Yeon Jeong ◽  
Hyung-Soon Yim ◽  
Ji-Young Ryu ◽  
Hyun Sook Lee ◽  
Jung-Hyun Lee ◽  
...  
Keyword(s):  
Functional Genomics ◽  
Dna Polymerase ◽  
Room Temperature ◽  
Cost Effective ◽  
Time Saving ◽  
Ligation Independent Cloning ◽  
Effective Time ◽  
Cloning Method ◽  
Directional Cloning ◽  
One Step

ABSTRACTWe developed one-step sequence- and ligation-independent cloning (SLIC) as a simple, cost-effective, time-saving, and versatile cloning method. Highly efficient and directional cloning can be achieved by direct bacterial transformation 2.5 min after mixing any linearized vector, an insert(s) prepared by PCR, and T4 DNA polymerase in a tube at room temperature.

scholarly journals One-step purification of Taq DNA polymerase using nucleotide-mimetic affinity chromatography

2007 ◽  
Vol 2 (1) ◽  
pp. 121-132 ◽  
Author(s):  
Sotirios Melissis ◽  
Nikolaos E. Labrou ◽  
Yannis D. Clonis
Keyword(s):  
Affinity Chromatography ◽  
Dna Polymerase ◽  
Taq Dna Polymerase ◽  
One Step

Preparation of long sticky ends for universal ligation-independent cloning: Sequential T4 DNA polymerase treatments

2009 ◽  
Vol 107 (6) ◽  
pp. 668-669 ◽  
Author(s):  
Akira Tachibana ◽  
Kazuo Tohiguchi ◽  
Takayuki Ueno ◽  
Yuichi Setogawa ◽  
Ayako Harada ◽  
...  
Keyword(s):  
Dna Polymerase ◽  
Ligation Independent Cloning ◽  
Sticky Ends

scholarly journals Development of an efficient one-step real-time reverse transcription polymerase chain reaction method for severe acute respiratory syndrome-coronavirus-2 detection

PLoS ONE ◽  
2021 ◽  
Vol 16 (6) ◽  
pp. e0252789
Author(s):  
Yukiko Nakura ◽  
Heng Ning Wu ◽  
Yuya Okamoto ◽  
Muneyuki Takeuchi ◽  
Koichiro Suzuki ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Reverse Transcriptase ◽  
Real Time ◽  
Dna Polymerase ◽  
Internal Control ◽  
Leukemia Virus ◽  
Rt Pcr ◽  
Thermotoga Petrophila ◽  
One Step ◽  
Thermostable Dna Polymerase

The general methods to detect the RNA of severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) in clinical diagnostic testing involve reverse transcriptases and thermostable DNA polymerases. In this study, we compared the detection of SARS-CoV-2 by a one-step real-time RT-PCR method using a heat-resistant reverse transcriptase variant MM4 from Moloney murine leukemia virus, two thermostable DNA polymerase variants with reverse transcriptase activity from Thermotoga petrophila K4 and Thermococcus kodakarensis KOD1, or a wild-type DNA polymerase from Thermus thermophilus M1. The highest performance was achieved by combining MM4 with the thermostable DNA polymerase from T. thermophilus M1. These enzymes efficiently amplified specific RNA using uracil-DNA glycosylase (UNG) to remove contamination and human RNase P RNA amplification as an internal control. The standard curve was obtained from 5 to 105 copies of synthetic RNA. The one-step real-time RT-PCR method’s sensitivity and specificity were 99.44% and 100%, respectively (n = 213), compared to those of a commercially available diagnostic kit. Therefore, our method will be useful for the accurate detection and quantification of SARS-CoV-2.

scholarly journals One-step RNA pathogen detection with reverse transcriptase activity of a mutated thermostable Thermus aquaticus DNA polymerase

2010 ◽  
Vol 5 (2) ◽  
pp. 224-231 ◽  
Author(s):  
Ramon Kranaster ◽  
Matthias Drum ◽  
Nicole Engel ◽  
Manfred Weidmann ◽  
Frank T. Hufert ◽  
...  
Keyword(s):  
Reverse Transcriptase ◽  
Dna Polymerase ◽  
Pathogen Detection ◽  
Reverse Transcriptase Activity ◽  
Thermus Aquaticus ◽  
Transcriptase Activity ◽  
One Step
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