Article
Biochemistry & Molecular Biology
Luisa B. Huber, Navpreet Kaur, Melanie Henkel, Virginie Marchand, Yuri Motorin, Ann E. Ehrenhofer-Murray, Andreas Marx
Summary: More than 170 posttranscriptional RNA modifications have been identified, including pseudouridine (psi) and queuosine (Q), which play important roles in translation regulation. Current detection methods rely on chemical treatment of RNA samples, but we have developed a RT-active DNA polymerase variant, RT-KTq I614Y, that can directly identify psi and Q sites without the need for chemical treatment. This method, combined with next-generation sequencing, provides a more efficient and accurate way to detect RNA modifications.
NUCLEIC ACIDS RESEARCH
(2023)
Article
Chemistry, Multidisciplinary
Zhaocheng Liu, Minxue Zheng, Zhaobang Liu
Summary: Automated Sanger sequencing technology, with its advantages of high precision and low cost, remains useful despite the emergence of next-generation sequencing platforms. Studies have shown that sequence errors can occur in PCR products due to the presence of stable hairpin structures in the template.
Article
Multidisciplinary Sciences
Nosaibah Samman, Khawlah Al-Muhalhil, Atef Nehdi
Summary: This study describes a simple and efficient method for the production of high-quality recombinant Taq DNA polymerase using heat denaturation and nickel affinity chromatography. The purified enzyme showed comparable activity to commercially available counterparts and was also treated with DNase to remove DNA contaminants.
JOURNAL OF KING SAUD UNIVERSITY SCIENCE
(2023)
Article
Biotechnology & Applied Microbiology
Xinjia Li, Binbin Chen, Wanyi Chen, Zhongji Pu, Xin Qi, Lirong Yang, Jianping Wu, Haoran Yu
Summary: Engineering Taq DNA polymerase (TaqPol) for improved activity, stability and sensitivity is crucial for its wide applications. Multiple sequence alignment (MSA) is used to design TaqPol mutations and two double mutations are generated. Mutations targeted on coevolutionary residues are applied to rescue stability or activity loss, and a small-scale MSA is used to predict single variants with increased stability, activity, and sensitivity.
APPLIED MICROBIOLOGY AND BIOTECHNOLOGY
(2023)
Article
Biochemistry & Molecular Biology
Sun Ju Lee, Sang-Yong Park, Kwang-Ho Lee, Min-Woo Lee, Chae-Yeon Yu, Jaeyoung Maeng, Hyeong-Dong Kim, Suhng Wook Kim
Summary: We developed a method using Escherichia coli that expresses thermostable Taq DNA polymerase directly in the PCR without purification. EcoliTaq showed activity comparable to that of commercial DNA polymerase and remained stable for 3 months. It proved to be useful as a DNA polymerase in hot-start PCR by effectively minimizing non-specific amplification.
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
(2023)
Article
Virology
Lakshminarayan M. Iyer, Vivek Anantharaman, Arunkumar Krishnan, A. Maxwell Burroughs, L. Aravind
Summary: Jumbo phages, with their extraordinary genome size, exhibit diverse adaptations across major biological systems and can be classified into three higher-order groups based on conserved gene patterns. Additionally, sensitive sequence analysis methods have identified novel protein-modifying enzymes that may help exploit host-machinery.
Article
Biotechnology & Applied Microbiology
Yongsong Zhou, Fan Bo, Tian Tian, Buling Wu, Bofeng Zhu
Summary: Due to the unique feature of Taq DNA polymerase, extra nucleotides can be added to the 3' terminus of PCR products, leading to the formation of an additional peak at the DYS391 locus. In this study, the formation mechanism of this artifact, known as excessive addition split peak (EASP), was explored through the analysis of PCR primers, amplicon sequences, and storage conditions. The EASP was found to be a result of 3' end non-template extension by Taq DNA polymerase, and it was influenced by primer sequences and PCR product storage conditions.
FRONTIERS IN BIOENGINEERING AND BIOTECHNOLOGY
(2023)
Article
Engineering, Environmental
Xinglu Jiang, Kangjing Li, Beibei Xie, Jun Zhou
Summary: By using a mixture of Taq DNA polymerase and BSA, a biomimetic enzyme hydrogel can be formed, held together mainly by noncovalent bonds. Temperature cycling disrupts the noncovalent bonds, releasing Taq DNA polymerase to synthesize DNA in a velocity-controlled manner. Differences in the initial DNA template can lead to variation in primer consumption, causing errors in qPCR Ct values. The slopes of the linear amplification curve can be used to calibrate Ct values and achieve more accurate detection of the initial content of the target DNA.
CHEMICAL ENGINEERING JOURNAL
(2023)
Article
Biotechnology & Applied Microbiology
Wayne M. Barnes, Zhian Zhang, Milko B. Kermekchiev
Summary: Changing an amino acid of Taq DNA polymerase leads to faster PCR; surprising results from negative controls reveal additional abilities of the enzyme.
FRONTIERS IN BIOENGINEERING AND BIOTECHNOLOGY
(2021)
Article
Nanoscience & Nanotechnology
Sneha Mittal, Milan Kumar Jena, Biswarup Pathak
Summary: This study explores the potential of using solid-state graphene nanogap electrodes for labeling DNA nucleotides in DNA sequencing. The results show that labeling nucleotides can enhance signal-to-noise ratio, control translocation velocity, and improve orientational variations. The proposed sequencing device exhibits high sensitivity and selectivity characteristics that are important for experimental purposes.
ACS APPLIED NANO MATERIALS
(2022)
Article
Biochemistry & Molecular Biology
Fumitaka Hashiya, Hirotaka Murase, Akash Chandela, Haruka Hiraoka, Masahito Inagaki, Yuko Nakashima, Naoko Abe, Mayu Nakamura, Goro Terai, Yasuaki Kimura, Kaori Ando, Natsuhisa Oka, Kiyoshi Asai, Hiroshi Abe
Summary: Controlling PCR fidelity is crucial in molecular biology, and high-fidelity PCR is essential for gene cloning. While most studies focus on protein engineering of polymerases for fidelity control, this report explores the use of chemically modified nucleotide substrates. The modification on P-gamma of nucleotide substrates was found to affect PCR fidelity, with deoxyadenosine tetraphosphate enhancing one mutation type and deoxyguanosine tetraphosphate suppressing another. By using excess amounts of deoxyguanosine tetraphosphate, both mutations were successfully suppressed, leading to improved overall fidelity.
Article
Chemistry, Analytical
Rui Zhang, Yuxuan Zhang, Li Ji, Lingli Gong, Yong Ji, Fengsheng Zhou, Ying Yin, Koukou Li, Ping Sun, Zhening Pu, Zhaocheng Liu, Jian Zou
Summary: Highly sensitive detection of gene variants is crucial for cancer diagnosis and treatment. In this study, a novel technique called HAC-PCR was presented, which can detect low-level DNA mutations with exceptional sensitivity. HAC-PCR achieves high sensitivity independently of strand-displacement, base modification, or the use of additional reagents, and introduces uniquely designed hairpin structures to improve the discrimination between wild-type and mutant targets. The optimized HAC-PCR consistently demonstrates the capability to discriminate mutant DNA at levels as low as 0.01% VAF for five mutation types.
MICROCHEMICAL JOURNAL
(2024)
Article
Biochemical Research Methods
Zhaocheng Liu, Xiushuai Li, Rui Zhang, Li Ji, Lingli Gong, Yong Ji, Fengsheng Zhou, Ying Yin, Koukou Li, Ping Sun, Zhening Pu, Qing Wang, Jian Zou
Summary: Detecting mutations related to tumors is crucial for cancer diagnosis and treatment. However, the presence of redundant wild DNA makes it a challenge to genotype low-copy mutant ctDNA. To address this, researchers have developed a new PCR strategy called UBC-PCR, which can identify low-abundance variants with high sensitivity, independent of strand displacement and base modification.
ANALYTICAL AND BIOANALYTICAL CHEMISTRY
(2023)
Article
Biotechnology & Applied Microbiology
Shi Rong Zhao, Bo Qu, Jae Kwang Kim, Bumkyu Lee
Summary: Polymerase chain reaction (PCR) is commonly used in diagnostic and molecular biology, but DNA contamination in Taq DNA polymerase reagents can lead to incorrect results, especially in monitoring GMOs. Investigation showed that many commercial Taq DNA polymerase reagents were contaminated with rRNA and antibiotic resistance genes, but decontamination was successful with nylon membrane treatment.
PLANT BIOTECHNOLOGY REPORTS
(2021)
Article
Entomology
Billy L. Amugune, Abneel K. Matharu, Paul Ouma, Francis Mutebi, Lynne Elson, Ulrike Fillinger, Juergen Kruecken
Summary: Tunga penetrans is an invasive flea that causes tungiasis, and a low-cost PCR-based tool for the identification of T. penetrans is needed to understand its ecology and monitor its development in soil.
Article
Microbiology
Shawn Walsh, David S. Peters, Peter A. Smith, Arryn Craney, Melissa M. Dix, Benjamin F. Cravatt, Floyd E. Romesberg
ANTIMICROBIAL AGENTS AND CHEMOTHERAPY
(2019)
Article
Chemistry, Multidisciplinary
Floyd E. Romesberg
ISRAEL JOURNAL OF CHEMISTRY
(2019)
Article
Biochemistry & Molecular Biology
Vivian T. Dien, Matthew Holcomb, Floyd E. Romesberg
Article
Biochemistry & Molecular Biology
Ramkrishna Adhikary, Jorg Zimmermann, Robyn L. Stanfield, Ian A. Wilson, Wayne Yu, Masayuki Oda, Floyd E. Romesberg
Article
Chemistry, Multidisciplinary
Aaron W. Feldman, Vivian T. Dien, Rebekah J. Karadeema, Emil C. Fischer, Yanbo You, Brooke A. Anderson, Ramanarayanan Krishnamurthy, Jason S. Chen, Lingjun Li, Floyd E. Romesberg
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
(2019)
Article
Chemistry, Multidisciplinary
Qian Shao, Tingjian Chen, Kai Sheng, Zhixia Liu, Zhuochen Zhang, Floyd E. Romesberg
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
(2020)
Article
Chemistry, Multidisciplinary
Anne Xiao-Zhou Zhou, Kai Sheng, Aaron W. Feldman, Floyd E. Romesberg
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
(2019)
Article
Biochemistry & Molecular Biology
Emil C. Fischer, Koji Hashimoto, Yorke Zhang, Aaron W. Feldman, Vivian T. Dien, Rebekah J. Karadeema, Ramkrishna Adhikary, Michael P. Ledbetter, Ramanarayanan Krishnamurthy, Floyd E. Romesberg
NATURE CHEMICAL BIOLOGY
(2020)
Article
Plant Sciences
Yun Xuan Tan, David S. Peters, Shawn Walsh, Matthew Holcomb, Diogo Santos-Martins, Stefano Forli, Floyd E. Romesberg
JOURNAL OF NATURAL PRODUCTS
(2020)
Review
Chemistry, Multidisciplinary
Miglena Manandhar, Eugene Chun, Floyd E. Romesberg
Summary: Almost all natural proteins consist of only 20 amino acids, but the ability to encode other amino acids opens up the possibility of producing proteins with novel functions, including more optimal therapeutic proteins. The field of genetic code expansion is moving towards commercialization for the production of improved protein therapeutics through academic and industrial progress.
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
(2021)
Article
Chemistry, Multidisciplinary
Koji Hashimoto, Emil C. Fischer, Floyd E. Romesberg
Summary: The researchers have developed semisynthetic organisms that can store and retrieve increased information by using a series of synthetic, unnatural base pairs. Their results demonstrate that the E. coli RNA polymerase can transcribe DNA containing unnatural base pairs, and these advancements should facilitate the creation of next generation SSOs.
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
(2021)
Review
Biology
Floyd E. Romesberg
Summary: Recent focus has been on developing proteins for human use, particularly in medicine. However, the limited functionality of natural proteins, made up of a limited number of canonical amino acids, makes it difficult to discover variants with desired functions. Expanding the genetic code by adding new codon/anticodon pairs to assign to non-canonical amino acids is expected to greatly improve the discovery of proteins with different functions. This article reviews over 20 years of effort leading to the discovery of synthetic nucleotides that can form unnatural base pairs and be incorporated into DNA for protein translation. It also discusses ongoing efforts to use these findings in the production of next-generation protein therapeutics by a pharmaceutical company.
PHILOSOPHICAL TRANSACTIONS OF THE ROYAL SOCIETY B-BIOLOGICAL SCIENCES
(2023)
Article
Chemistry, Multidisciplinary
Anne X-Z Zhou, Xiyu Dong, Floyd E. Romesberg
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
(2020)
Meeting Abstract
Chemistry, Multidisciplinary
Floyd Romesberg
ABSTRACTS OF PAPERS OF THE AMERICAN CHEMICAL SOCIETY
(2018)
Review
Chemistry, Multidisciplinary
Aaron W. Feldmann, Floyd E. Romesberg
ACCOUNTS OF CHEMICAL RESEARCH
(2018)
