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Journal articles on the topic 'Phi29 DNA Polymerase'

Author: Grafiati

Published: 20 October 2023

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1

del Prado, Santos, Lázaro, Salas, and de Vega. "The Loop of the TPR1 Subdomain of Phi29 DNA Polymerase Plays a Pivotal Role in Primer-Terminus Stabilization at the Polymerization Active Site." Biomolecules 9, no. 11 (October 24, 2019): 648. http://dx.doi.org/10.3390/biom9110648.

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Bacteriophage Phi29 DNA polymerase belongs to the protein-primed subgroup of family B DNA polymerases that use a terminal protein (TP) as a primer to initiate genome replication. The resolution of the crystallographic structure showed that it consists of an N-terminal domain with the exonuclease activity and a C-terminal polymerization domain. It also has two subdomains specific of the protein-primed DNA polymerases; the TP Regions 1 (TPR1) that interacts with TP and DNA, and 2 (TPR2), that couples both processivity and strand displacement to the enzyme. The superimposition of the structures of the apo polymerase and the polymerase in the polymerase/TP heterodimer shows that the structural changes are restricted almost to the TPR1 loop (residues 304–314). In order to study the role of this loop in binding the DNA and the TP, we changed the residues Arg306, Arg308, Phe309, Tyr310, and Lys311 into alanine, and also made the deletion mutant Δ6 lacking residues Arg306–Lys311. The results show a defective TP binding capacity in mutants R306A, F309A, Y310A, and Δ6. The additional impaired primer-terminus stabilization at the polymerization active site in mutants Y310A and Δ6 allows us to propose a role for the Phi29 DNA polymerase TPR1 loop in the proper positioning of the DNA and TP-priming 3’-OH termini at the preinsertion site of the polymerase to enable efficient initiation and further elongation steps during Phi29 TP-DNA replication.

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2

Sakatani, Yoshihiro, Ryo Mizuuchi, and Norikazu Ichihashi. "In vitro evolution of phi29 DNA polymerases through compartmentalized gene expression and rolling-circle replication." Protein Engineering, Design and Selection 32, no. 11 (November 2019): 481–87. http://dx.doi.org/10.1093/protein/gzaa011.

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Abstract Phi29 DNA polymerase is widely used for DNA amplification through rolling-circle replication or multiple displacement amplification. Here, we performed completely in vitro artificial evolution of phi29 DNA polymerase by combining the in vitro compartmentalization and the gene expression-coupled rolling-circle replication of a circular DNA encoding the polymerase. We conducted the experiments in six different conditions composed of three different levels of inhibitor concentrations with two different DNA labeling methods. One of the experiments was performed in our previous study and the other five experiments were newly conducted in this study. Under all conditions, we found several mutations that enhance the rolling-circle amplification by the polymerase when it was expressed in the reconstituted gene expression system. Especially, a combinatorial mutant polymerase (K555T/D570N) exhibits significantly higher rolling-circle activity than the wild type. These highly active mutant polymerases would be useful for various applications.

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3

Kamtekar, Satwik. "Phi29 DNA polymerase: structure and sequencing." Acta Crystallographica Section A Foundations and Advances 75, a1 (July 20, 2019): a139. http://dx.doi.org/10.1107/s010876731909860x.

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4

Krzywkowski, Tomasz, Malte Kühnemund, Di Wu, and Mats Nilsson. "Limited reverse transcriptase activity of phi29 DNA polymerase." Nucleic Acids Research 46, no. 7 (March 15, 2018): 3625–32. http://dx.doi.org/10.1093/nar/gky190.

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5

Tenaglia, Enrico, Yuki Imaizumi, Yuji Miyahara, and Carlotta Guiducci. "Isothermal multiple displacement amplification of DNA templates in minimally buffered conditions using phi29 polymerase." Chemical Communications 54, no. 17 (2018): 2158–61. http://dx.doi.org/10.1039/c7cc09609g.

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6

Torres, Leticia L., and Vitor B. Pinheiro. "Xenobiotic Nucleic Acid (XNA) Synthesis by Phi29 DNA Polymerase." Current Protocols in Chemical Biology 10, no. 2 (May 18, 2018): e41. http://dx.doi.org/10.1002/cpch.41.

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7

Li, Shasha, Su Liu, Yicheng Xu, Rufeng Zhang, Yihan Zhao, Xiaonan Qu, Yu Wang, Jiadong Huang, and Jinghua Yu. "Robust and highly specific fluorescence sensing of Salmonella typhimurium based on dual-functional phi29 DNA polymerase-mediated isothermal circular strand displacement polymerization." Analyst 144, no. 16 (2019): 4795–802. http://dx.doi.org/10.1039/c9an00843h.

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A simple and robust fluorescence sensing strategy has been developed for the detection of pathogenic bacteria by the combination of the dual functionality of phi29 DNA polymerase with isothermal circular strand displacement polymerization (ICSDP).

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8

Xu, Yun, Simon Gao, John F. Bruno, Benjamin J. Luft, and John J. Dunn. "Rapid detection and identification of a pathogen’s DNA using Phi29 DNA polymerase." Biochemical and Biophysical Research Communications 375, no. 4 (October 2008): 522–25. http://dx.doi.org/10.1016/j.bbrc.2008.08.082.

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9

Johne, Reimar, Hermann Müller, Annabel Rector, Marc van Ranst, and Hans Stevens. "Rolling-circle amplification of viral DNA genomes using phi29 polymerase." Trends in Microbiology 17, no. 5 (May 2009): 205–11. http://dx.doi.org/10.1016/j.tim.2009.02.004.

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10

Kesici, Merve-Zeynep, Philip Tinnefeld, and Andrés Manuel Vera. "A simple and general approach to generate photoactivatable DNA processing enzymes." Nucleic Acids Research 50, no. 6 (December 14, 2021): e31-e31. http://dx.doi.org/10.1093/nar/gkab1212.

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Abstract DNA processing enzymes, such as DNA polymerases and endonucleases, have found many applications in biotechnology, molecular diagnostics, and synthetic biology, among others. The development of enzymes with controllable activity, such as hot-start or light-activatable versions, has boosted their applications and improved the sensitivity and specificity of the existing ones. However, current approaches to produce controllable enzymes are experimentally demanding to develop and case-specific. Here, we introduce a simple and general method to design light-start DNA processing enzymes. In order to prove its versatility, we applied our method to three DNA polymerases commonly used in biotechnology, including the Phi29 (mesophilic), Taq, and Pfu polymerases, and one restriction enzyme. Light-start enzymes showed suppressed polymerase, exonuclease, and endonuclease activity until they were re-activated by an UV pulse. Finally, we applied our enzymes to common molecular biology assays and showed comparable performance to commercial hot-start enzymes.

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11

Lieberman, Kate R., Gerald M. Cherf, Michael J. Doody, Felix Olasagasti, Yvette Kolodji, and Mark Akeson. "Processive Replication of Single DNA Molecules in a Nanopore Catalyzed by phi29 DNA Polymerase." Journal of the American Chemical Society 132, no. 50 (December 22, 2010): 17961–72. http://dx.doi.org/10.1021/ja1087612.

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12

Taniguchi, R., C. Masaki, Y. Murashima, M. Makino, T. Kojo, T. Nakamoto, and R. Hosokawa. "DNA amplification using phi29 DNA polymerase validates gene polymorphism analysis from buccal mucosa samples." Journal of Prosthodontic Research 55, no. 3 (July 2011): 165–70. http://dx.doi.org/10.1016/j.jpor.2010.12.001.

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13

Povilaitis, Tadas, Gediminas Alzbutas, Rasa Sukackaite, Juozas Siurkus, and Remigijus Skirgaila. "In vitroevolution of phi29 DNA polymerase using isothermal compartmentalized self replication technique." Protein Engineering, Design and Selection 29, no. 12 (November 2016): 617–28. http://dx.doi.org/10.1093/protein/gzw052.

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14

Berman, Andrea J., Satwik Kamtekar, Jessica L. Goodman, José M. Lázaro, Miguel de Vega, Luis Blanco, Margarita Salas, and Thomas A. Steitz. "Structures of phi29 DNA polymerase complexed with substrate: the mechanism of translocation in B-family polymerases." EMBO Journal 26, no. 14 (July 5, 2007): 3494–505. http://dx.doi.org/10.1038/sj.emboj.7601780.

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15

Manrao, Elizabeth A., Ian M. Derrington, Andrew H. Laszlo, Kyle W. Langford, Matthew K. Hopper, Nathaniel Gillgren, Mikhail Pavlenok, Michael Niederweis, and Jens H. Gundlach. "Reading DNA at single-nucleotide resolution with a mutant MspA nanopore and phi29 DNA polymerase." Nature Biotechnology 30, no. 4 (March 25, 2012): 349–53. http://dx.doi.org/10.1038/nbt.2171.

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16

Dean, F. B. "Rapid Amplification of Plasmid and Phage DNA Using Phi29 DNA Polymerase and Multiply-Primed Rolling Circle Amplification." Genome Research 11, no. 6 (June 1, 2001): 1095–99. http://dx.doi.org/10.1101/gr.180501.

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17

Kim, Subin, and Ja Yil Lee. "Study on biophysical properties of Phi29 DNA polymerase using a novel single-molecule imaging technique DNA curtain." Biophysical Journal 122, no. 3 (February 2023): 356a. http://dx.doi.org/10.1016/j.bpj.2022.11.1972.

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18

Liang, Jingjing, Jiaqi Zhou, Jianxi Tan, Zefeng Wang, and Le Deng. "Aptamer-Based Fluorescent Determination of Salmonella paratyphi A Using Phi29-DNA Polymerase-Assisted Cyclic Amplification." Analytical Letters 52, no. 6 (September 20, 2018): 919–31. http://dx.doi.org/10.1080/00032719.2018.1505901.

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19

Sato, M. "Repeated GenomiPhi, phi29 DNA polymerase-based rolling circle amplification, is useful for generation of large amounts of plasmid DNA." Nucleic Acids Symposium Series 48, no. 1 (November 1, 2004): 147–48. http://dx.doi.org/10.1093/nass/48.1.147.

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20

Gao, Yaping, Yun He, Liyi Chen, Xing Liu, Igor Ivanov, Xuerui Yang, and Hui Tian. "Chimeric Phi29 DNA polymerase with helix–hairpin–helix motifs shows enhanced salt tolerance and replication performance." Microbial Biotechnology 14, no. 4 (May 19, 2021): 1642–56. http://dx.doi.org/10.1111/1751-7915.13830.

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21

Lagunavicius, A., Z. Kiveryte, V. Zimbaite-Ruskuliene, T. Radzvilavicius, and A. Janulaitis. "Duality of polynucleotide substrates for Phi29 DNA polymerase: 3'->5' RNase activity of the enzyme." RNA 14, no. 3 (January 18, 2008): 503–13. http://dx.doi.org/10.1261/rna.622108.

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22

Chen, Anyi, Guo-Feng Gui, Ying Zhuo, Ya-Qin Chai, Yun Xiang, and Ruo Yuan. "Signal-off Electrochemiluminescence Biosensor Based on Phi29 DNA Polymerase Mediated Strand Displacement Amplification for MicroRNA Detection." Analytical Chemistry 87, no. 12 (May 22, 2015): 6328–34. http://dx.doi.org/10.1021/acs.analchem.5b01168.

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23

Niel, Christian, Leonardo Diniz-Mendes, and Sylvie Devalle. "Rolling-circle amplification of Torque teno virus (TTV) complete genomes from human and swine sera and identification of a novel swine TTV genogroup." Journal of General Virology 86, no. 5 (May 1, 2005): 1343–47. http://dx.doi.org/10.1099/vir.0.80794-0.

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Multiply primed rolling-circle amplification is a novel technology that uses bacteriophage phi29 DNA polymerase to amplify circular DNA molecules, without the need for prior knowledge of their sequences. In an attempt to detect Torque teno virus (TTV), rolling-circle amplification was used to amplify DNA extracted from eight human and four pig serum samples. All samples gave high molecular weight (>30 kb) amplification products. By restriction endonuclease digestion, these products generated DNA fragments whose sizes were consistent with those of human TTV (3·8 kb) and swine TTV (Sd-TTV; 2·9 kb) genomes. Two TTV isolates derived from a single AIDS patient, as well as two Sd-TTV isolates derived from a single pig, were characterized by complete nucleotide sequencing. One of the Sd-TTV isolates showed very low (43–45 %) nucleotide sequence similarity to the other Sd-TTV isolate and to the prototype isolate Sd-TTV31, and could be considered the prototype of a novel genogroup.

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24

Wang, Yuhan, Jiaxuan Xiao, Xiaona Lin, Amira Waheed, Ayyanu Ravikumar, Zhen Zhang, Yanmin Zou, and Chengshui Chen. "A Self-Assembled G-Quadruplex/Hemin DNAzyme-Driven DNA Walker Strategy for Sensitive and Rapid Detection of Lead Ions Based on Rolling Circle Amplification." Biosensors 13, no. 8 (July 26, 2023): 761. http://dx.doi.org/10.3390/bios13080761.

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Herein, a sensitive biosensor is constructed based on a novel rolling circle amplification (RCA) for colorimetric quantification of lead ion (Pb2+). At the detection system, GR5 DNAzymes are modified on the surface of an immunomagnetic bead, and Pb2+ is captured by the aptamer, inducing the disintegration of the GR5 DNAzyme and the release of the DNA walker. After the introduction of the template DNA, T4 DNA ligase, and phi29 DNA polymerase, an RCA is initiated for the sensitivity improvement of this method. Moreover, a G4-hemin DNAzyme is formed as a colorimetric signal, owing to its peroxide-like activity to catalyze the TMB-H2O2 substrate. Under the optimized conditions, the limit of detection (LOD) of this fabricated biosensor could reach 3.3 pM for Pb2+ with a concentration in the range of 0.01–1000 nM. Furthermore, the results of real samples analysis demonstrate its satisfactory accuracy, implying its great potential in the rapid detection of heavy metals in the environment.

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25

Truniger, V. "A positively charged residue of phi29 DNA polymerase, highly conserved in DNA polymerases from families A and B, is involved in binding the incoming nucleotide." Nucleic Acids Research 30, no. 7 (April 1, 2002): 1483–92. http://dx.doi.org/10.1093/nar/30.7.1483.

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26

Zhu, Qiang, Ting Fang, Yijun Zhou, Yiwen Yang, Yueyan Cao, Qiuyue Wang, Yuguo Huang, et al. "Effect of phi29 polymerase-based multiple strand displacement whole genome amplification on the proportion in DNA mixtures." Forensic Science International: Genetics Supplement Series 7, no. 1 (December 2019): 841–42. http://dx.doi.org/10.1016/j.fsigss.2019.10.197.

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27

Gadkar, Vijay, and Matthias C. Rillig. "Application of Phi29 DNA polymerase mediated whole genome amplification on single spores of arbuscular mycorrhizal (AM) fungi." FEMS Microbiology Letters 242, no. 1 (January 2005): 65–71. http://dx.doi.org/10.1016/j.femsle.2004.10.041.

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28

Million, Matthieu, Maxime Gaudin, Cléa Melenotte, Lionel Chasson, Sophie Edouard, Constance Verdonk, Elsa Prudent, et al. "Metagenomic Analysis of Microdissected Valvular Tissue for Etiological Diagnosis of Blood Culture–Negative Endocarditis." Clinical Infectious Diseases 70, no. 11 (July 15, 2019): 2405–12. http://dx.doi.org/10.1093/cid/ciz655.

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Abstract Background Etiological diagnosis is a key to therapeutic adaptation and improved prognosis, particularly for infections such as endocarditis. In blood culture–negative endocarditis (BCNE), 22% of cases remain undiagnosed despite an updated comprehensive syndromic approach. This prompted us to develop a new diagnostic approach. Methods Eleven valves from 10 BCNE patients were analyzed using a method that combines human RNA bait-depletion with phi29 DNA polymerase-based multiple displacement amplification and shotgun DNA sequencing. An additional case in which a microbe was serendipitously visualized by immunofluorescence was analyzed using the same method, but after laser capture microdissection. Results Background DNA prevented any diagnosis in cases analyzed without microdissection because the majority of sequences were contaminants. Moraxella sequences were dramatically enriched in the stained microdissected region of the additional case. A consensus genome sequence of 2.4 Mbp covering more than 94% of the Moraxella osloensis KSH reference genome was reconstructed with 234X average coverage. Several antibiotic-resistance genes were observed. Etiological diagnosis was confirmed using Western blot and specific polymerase chain reaction with sequencing on a different valve sample. Conclusions Microdissection could be a key to the metagenomic diagnosis of infectious diseases when a microbe is visualized but remains unidentified despite an updated optimal approach. Moraxella osloensis should be tested in blood culture–negative endocarditis.

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29

Takahashi, Hirokazu, Hiroyuki Yamazaki, Satoshi Akanuma, Hiroko Kanahara, Toshiyuki Saito, Tomoyuki Chimuro, Takayoshi Kobayashi, et al. "Preparation of Phi29 DNA Polymerase Free of Amplifiable DNA Using Ethidium Monoazide, an Ultraviolet-Free Light-Emitting Diode Lamp and Trehalose." PLoS ONE 9, no. 2 (February 5, 2014): e82624. http://dx.doi.org/10.1371/journal.pone.0082624.

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30

Sato, Masahiro, Masato Ohtsuka, and Yuhsuke Ohmi. "Usefulness of repeated GenomiPhi, a phi29 DNA polymerase-based rolling circle amplification kit, for generation of large amounts of plasmid DNA." Biomolecular Engineering 22, no. 4 (October 2005): 129–32. http://dx.doi.org/10.1016/j.bioeng.2005.05.001.

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31

de Vega, M., J. M. Lazaro, M. Salas, and L. Blanco. "Primer-terminus stabilization at the 3′-5′ exonuclease active site of phi29 DNA polymerase. Involvement of two amino acid residues highly conserved in proofreading DNA polymerases." EMBO Journal 15, no. 5 (March 1996): 1182–92. http://dx.doi.org/10.1002/j.1460-2075.1996.tb00457.x.

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32

Kim, Min-Soo, Eun-Jin Park, Seong Woon Roh, and Jin-Woo Bae. "Diversity and Abundance of Single-Stranded DNA Viruses in Human Feces." Applied and Environmental Microbiology 77, no. 22 (September 23, 2011): 8062–70. http://dx.doi.org/10.1128/aem.06331-11.

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ABSTRACTIn this study, we investigated the abundance and diversity of single-stranded DNA (ssDNA) viruses in fecal samples from five healthy individuals through a combination of serial filtration and CsCl gradient ultracentrifugation. Virus abundance ranged from 108to 109per gram of feces, and virus-to-bacterium ratios were much lower (less than 0.1) than those observed in aquatic environments (5 to 10). Viral DNA was extracted and randomly amplified using phi29 polymerase and analyzed through high-throughput 454 pyrosequencing. Among 400,133 sequences, an average of 86.2% viromes were previously uncharacterized in public databases. Among previously known viruses, double-stranded DNA podophages (52 to 74%), siphophages (11 to 30%), myophages (1 to 4%), and ssDNA microphages (3 to 9%) were major constituents of human fecal viromes. A phylogenetic analysis of 24 large contigs of microphages based on conserved capsid protein sequences revealed five distinct newly discovered evolutionary microphage groups that were distantly related to previously known microphages. Moreover, putative capsid protein sequences of five contigs were closely related to prophage-like sequences in the genomes of threeBacteroidesand threePrevotellastrains, suggesting thatBacteroidesandPrevotellaare the sources of infecting microphages in their hosts.

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33

Sakatani, Yoshihiro, Norikazu Ichihashi, and Tetsuya Yomo. "2P262 Establishment of a self-replication system using phi29 DNA polymerase(20. Origin of life & Evolution,Poster)." Seibutsu Butsuri 54, supplement1-2 (2014): S238. http://dx.doi.org/10.2142/biophys.54.s238_4.

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34

Ye, Yan, Yao Lin, Zilin Chi, Jiasheng Zhang, Fan Cai, Youzhi Zhu, Dianping Tang, and Qingqiang Lin. "Rolling circle amplification (RCA) -based biosensor system for the fluorescent detection of miR-129-2-3p miRNA." PeerJ 10 (October 24, 2022): e14257. http://dx.doi.org/10.7717/peerj.14257.

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Herein, a versatile fluorescent bioanalysis platform for sensitive and specific screening of target miRNA (miR-129-2-3p) was innovatively designed by applying target-induced rolling circle amplification (RCA) for efficient signal amplification. Specifically, miR-129-2-3p was used as a ligation template to facilitate its ligation with padlock probes, followed by an RCA reaction in the presence of phi29 DNA polymerase. The dsDNA fragments and products were stained by SYBR Green I and then detected by fluorescence spectrophotometry. As a result, miR-129-2-3p concentrations as low as 50 nM could be detected. Furthermore, the expression of miR-129-2-3p in breast cancer patients was about twice that in healthy people. Therefore, the results indicated that the RCA-based biosensor system could be a valuable platform for miRNA detection in clinical diagnosis and biomedical study.

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35

Wu, Bingyun, Hiroyuki Kurokochi, and Taizo Hogetsu. "Development of 12 microsatellite markers in Euptelea polyandra by a random tailed genome-walking method using Phi29 DNA polymerase." Conservation Genetics Resources 1, no. 1 (May 19, 2009): 59–61. http://dx.doi.org/10.1007/s12686-009-9014-y.

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36

Taniguchi, Ryoji, Chihiro Masaki, Yuhi Murashima, Michiko Makino, Tatsuro Kojo, Tetsuji Nakamoto, and Ryuji Hosokawa. "Erratum to “DNA amplification using phi29 DNA polymerase validates gene polymorphism analysis from buccal mucosa samples” [J. Prosthodont. Res. 55 (2011) 165–170]." Journal of Prosthodontic Research 55, no. 4 (October 2011): 266. http://dx.doi.org/10.1016/j.jpor.2011.08.001.

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37

Lagunavicius, A., E. Merkiene, Z. Kiveryte, A. Savaneviciute, V. Zimbaite-Ruskuliene, T. Radzvilavicius, and A. Janulaitis. "Novel application of Phi29 DNA polymerase: RNA detection and analysis in vitro and in situ by target RNA-primed RCA." RNA 15, no. 5 (March 24, 2009): 765–71. http://dx.doi.org/10.1261/rna.1279909.

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38

Pan, Xinghua, Alexander Eckehart Urban, Dean Palejev, Vincent Schulz, Fabian Grubert, Yiping Hu, Michael Snyder, and Sherman M. Weissman. "A procedure for highly specific, sensitive, and unbiased whole-genome amplification." Proceedings of the National Academy of Sciences 105, no. 40 (October 1, 2008): 15499–504. http://dx.doi.org/10.1073/pnas.0808028105.

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Highly specific amplification of complex DNA pools without bias or template-independent products (TIPs) remains a challenge. We have developed a method using phi29 DNA polymerase and trehalose and optimized control of amplification to create micrograms of specific amplicons without TIPs from down to subfemtograms of DNA. With an input of as little as 0.5–2.5 ng of human gDNA or a few cells, the product could be close to native DNA in locus representation. The amplicons from 5 and 0.5 ng of DNA faithfully demonstrated all previously known heterozygous segmental duplications and deletions (3 Mb to 18 kb) located on chromosome 22 and even a homozygous deletion smaller than 1 kb with high-resolution chromosome-wide comparative genomic hybridization. With 550k Infinium BeadChip SNP typing, the >99.7% accuracy was compared favorably with results on unamplified DNA. Importantly, underrepresentation of chromosome termini that occurred with GenomiPhi v2 was greatly rescued with the present procedure, and the call rate and accuracy of SNP typing were also improved for the amplicons with a 0.5-ng, partially degraded DNA input. In addition, the amplification proceeded logarithmically in terms of total yield before saturation; the intact cells was amplified >50 times more efficiently than an equivalent amount of extracted DNA; and the locus imbalance for amplicons with 0.1 ng or lower input of DNA was variable, whereas for higher input it was largely reproducible. This procedure facilitates genomic analysis with single cells or other traces of DNA, and generates products suitable for analysis by massively parallel sequencing as well as microarray hybridization.

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39

Alsmadi, Osama, Fadi Alkayal, Dorota Monies, and Brian F. Meyer. "Specific and complete human genome amplification with improved yield achieved by phi29 DNA polymerase and a novel primer at elevated temperature." BMC Research Notes 2, no. 1 (2009): 48. http://dx.doi.org/10.1186/1756-0500-2-48.

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40

Eisenbrandt, R. "Phi29 DNA polymerase residues Tyr59, His61 and Phe69 of the highly conserved ExoII motif are essential for interaction with the terminal protein." Nucleic Acids Research 30, no. 6 (March 15, 2002): 1379–86. http://dx.doi.org/10.1093/nar/30.6.1379.

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41

Knierim, D., and E. Maiss. "Application of Phi29 DNA polymerase in identification and full-length clone inoculation of tomato yellow leaf curl Thailand virus and tobacco leaf curl Thailand virus." Archives of Virology 152, no. 5 (January 18, 2007): 941–54. http://dx.doi.org/10.1007/s00705-006-0914-9.

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42

Lu, Na, Junji Li, Changwei Bi, Jing Guo, Yuhan Tao, Kaihao Luan, Jing Tu, and Zuhong Lu. "ChimeraMiner: An Improved Chimeric Read Detection Pipeline and Its Application in Single Cell Sequencing." International Journal of Molecular Sciences 20, no. 8 (April 21, 2019): 1953. http://dx.doi.org/10.3390/ijms20081953.

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As the most widely-used single cell whole genome amplification (WGA) approach, multiple displacement amplification (MDA) has a superior performance, due to the high-fidelity and processivity of phi29 DNA polymerase. However, chimeric reads, generated in MDA, cause severe disruption in many single-cell studies. Herein, we constructed ChimeraMiner, an improved chimeric read detection pipeline for analyzing the sequencing data of MDA and classified the chimeric sequences. Two datasets (MDA1 and MDA2) were used for evaluating and comparing the efficiency of ChimeraMiner and previous pipeline. Under the same hardware condition, ChimeraMiner spent only 43.4% (43.8% for MDA1 and 43.0% for MDA2) processing time. Respectively, 24.4 million (6.31%) read pairs out of 773 million reads, and 17.5 million (6.62%) read pairs out of 528 million reads were accurately classified as chimeras by ChimeraMiner. In addition to finding 83.60% (17,639,371) chimeras, which were detected by previous pipelines, ChimeraMiner screened 6,736,168 novel chimeras, most of which were missed by the previous pipeline. Applying in single-cell datasets, all three types of chimera were discovered in each dataset, which introduced plenty of false positives in structural variation (SV) detection. The identification and filtration of chimeras by ChimeraMiner removed most of the false positive SVs (83.8%). ChimeraMiner revealed improved efficiency in discovering chimeric reads, and is promising to be widely used in single-cell sequencing.

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43

Jung, S., M. Reichenbach, R. Fries, E. Wolf, C. Gschoederer, J. Scherzer, T. Grupp, and H. D. Reichenbach. "316 GENOMIC EVALUATION OF BOVINE EMBRYOS WITHIN 24 HOURS." Reproduction, Fertility and Development 27, no. 1 (2015): 247. http://dx.doi.org/10.1071/rdv27n1ab316.

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The aim of this study was to develop a reliable procedure for genomic evaluation of bovine embryos to determine gender, polled status, and hereditary defects within 24 h after collection. German Simmental animals (n = 15) were superovulated (n = 25) using a standard protocol. Embryos were recovered on Day 7 (Day 0 = oestrus). A total of 217 embryos (morula, n = 130; early blastocyst, n = 43; blastocyst, n = 44) were biopsied with a steel blade attached to a micromanipulator. Biopsied cells were immediately transferred into 1 µL TE buffer to a 500 µL reaction tube and embryos were in vitro cultured until genomic results were available. For commonly used molecular genetic methods (e.g. 5′-exonuclease genotyping, PCR or high density genotyping) DNA amounts of 2–200 ng are required. However, the DNA quantity of a single diploid cell amounts to 6 pg only. The embryo biopsies used, usually consists of 10–30 cells, necessitating an artificial amplification of the embryonic genome. Taking all vital measures to avoid external DNA contamination, isothermal whole genome amplification was performed with the REPLI-g Mini Kit (Qiagen, Valencia, CA, USA) using random hexamers and Phi29-Polymerase. Depending on the number of cells, a total DNA amount of 4–7 µg was achieved. Polled status and gender was determined using PCR with subsequent gel-electrophoresis. 5′-exonuclease assays were used to obtain genotypes for the detection of genetic defects. At present, eight, mostly Simmental-specific genetic disorders can be examined: three traits associated with severe growth retardation, dwarfism (DW), Braunvieh-haplotype 2 (BH2) and stunted growth (FH2), the lethal skin disorder zinc deficiency-like syndrome (ZDL), a fertility trait bovine male subfertility (BMS), embryonic death Fleckvieh-haplotype 4 (FH4), a bleeding disorder thrombopathia (TP) and arachnomelia (A), within 24 h. On average, 8.7 embryos were biopsied per embryo recovery, i.e. 93.9% of the total number of transferable embryos. Fourteen embryo samples (6.5%) totally failed during analysis, possibly due to the loss of samples. In successful analyses, gender was undetermined in two embryos; remaining embryos were 52.2% female and 47.8% male. Polled status could be analysed in 92.6% of the embryos. The analyses of embryos for possible inherited genetic disorders (healthy, heterozygote, or homozygote; n = 578) were successful in 90.1%. The transfer of biopsied embryos (n = 30) led to 63.3% pregnancies (Day 42). A validation of the present results has to be done as soon as the produced calves are born, demonstrating the reliability of the procedure.Research was funded by the Bayerische Forschungsstiftung (AZ-1031-12).

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"5198543 Phi29 DNA polymerase." Biotechnology Advances 12, no. 1 (January 1994): 127. http://dx.doi.org/10.1016/0734-9750(94)90402-2.

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"5001050 PH phi29 DNA polymerase." Biotechnology Advances 9, no. 3 (January 1991): 445. http://dx.doi.org/10.1016/0734-9750(91)90880-5.

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Zhang, Jia, Xiaolu Su, Yefei Wang, Xiaohang Wang, Shiqi Zhou, Hui Jia, Xiaoyan Jing, Yanhai Gong, Jichao Wang, and Jian Xu. "Improved single-cell genome amplification by a high-efficiency phi29 DNA polymerase." Frontiers in Bioengineering and Biotechnology 11 (June 29, 2023). http://dx.doi.org/10.3389/fbioe.2023.1233856.

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Single-cell genomic whole genome amplification (WGA) is a crucial step in single-cell sequencing, yet its low amplification efficiency, incomplete and uneven genome amplification still hinder the throughput and efficiency of single-cell sequencing workflows. Here we introduce a process called Improved Single-cell Genome Amplification (iSGA), in which the whole single-cell sequencing cycle is completed in a high-efficient and high-coverage manner, through phi29 DNA polymerase engineering and process engineering. By establishing a disulfide bond of F137C-A377C, the amplification ability of the enzyme was improved to that of single-cell. By further protein engineering and process engineering, a supreme enzyme named HotJa Phi29 DNA Polymerase was developed and showed significantly better coverage (99.75%) at a higher temperature (40°C). High single-cell genome amplification ability and high coverage (93.59%) were also achieved for commercial probiotic samples. iSGA is more efficient and robust than the wild-type phi29 DNA polymerase, and it is 2.03-fold more efficient and 10.89-fold cheaper than the commercial Thermo Scientific EquiPhi29 DNA Polymerase. These advantages promise its broad applications in large-scale single-cell sequencing.

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Nelson, John R. "Random‐Primed, Phi29 DNA Polymerase‐Based Whole Genome Amplification." Current Protocols in Molecular Biology 105, no. 1 (January 2014). http://dx.doi.org/10.1002/0471142727.mb1513s105.

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Zhang, Jia, Xiaolu Su, Yefei Wang, Xiaohang Wang, Shiqi Zhou, Hui Jia, Xiaoyan Jing, Yanhai Gong, Jichao Wang, and Jian Xu. "Corrigendum: Improved single-cell genome amplification by a high-efficiency phi29 DNA polymerase." Frontiers in Bioengineering and Biotechnology 11 (August 28, 2023). http://dx.doi.org/10.3389/fbioe.2023.1263634.

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Zhang, Xi, Jingjing Chen, Pengfei Jiang, Heling Xu, Qi Zhang, Huan Zhang, Xiaohu Han, and Zeliang Chen. "A Phi29-based unbiased exponential amplification and genotyping approach improves pathogen detection in tick samples." Frontiers in Veterinary Science 9 (November 7, 2022). http://dx.doi.org/10.3389/fvets.2022.1025911.

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Ticks are vectors for many infectious diseases, such as spotted fever group (SFG) rickettsioses and borreliosis, and are valuable in the study of pathogen ecology. Ticks have several growth stages that vary considerably in size; therefore, in most cases, DNA extracted from ticks is insufficient for subsequent studies, particularly for multiple pathogen screening and genotyping. Unbiased amplification of DNA from tick samples before analysis is a major requirement for subsequent ecological surveys and other studies. Phi29 DNA polymerase, an enzyme that exhibits strand displacement activity, can exponentially amplify DNA randomly, generating large quantities of DNA. In the present study, we developed a Phi29-based unbiased exponential amplification (PEA) assay to obtain sufficient tick DNA for genetic analysis. By using tick-borne pathogen detection and genotyping as a model, we tested and evaluated the feasibility of the assay. DNA was extracted from single ticks and subjected to PEA. The results showed that tick DNA could be amplified up to 105 fold. The amplified products were successfully used for pathogen screening and genotyping. Rickettsia was successfully detected and genotyped in samples with amplified DNA from single ticks. Furthermore, we identified a new genotype of Rickettsia from ticks collected from Dandong city, Liaoning province, Northeast China. This PEA assay is universal and can be extended to other applications where the quantity of DNA is greatly limited.

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Tsuruta, Haruka, Yuina Sonohara, Kosuke Tohashi, Narumi Aoki Shioi, Shigenori Iwai, and Isao Kuraoka. "Effects of acetaldehyde-induced DNA lesions on DNA metabolism." Genes and Environment 42, no. 1 (January 6, 2020). http://dx.doi.org/10.1186/s41021-019-0142-7.

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Abstract Background Acetaldehyde, produced upon exposure to alcohol, cigarette smoke, polluted air and sugar, is a highly reactive compound that is carcinogenic to humans and causes a variety of DNA lesions in living human cells. Previously, we reported that acetaldehyde reacts with adjacent deoxyguanosine residues on oligonucleotides, but not with single deoxyguanosine residues or other deoxyadenosine, deoxycytosine, or thymidine residues, and revealed that it forms reversible intrastrand crosslinks with the dGpdG sequence (GG dimer). Results Here, we show that restriction enzymes that recognize a GG sequence digested acetaldehyde-treated plasmid DNA with low but significant efficiencies, whereas restriction enzymes that recognize other sequences were able to digest such DNA. This suggested that acetaldehyde produced GG dimers in plasmid DNA. Additionally, acetaldehyde-treated oligonucleotides were efficient in preventing digestion by the exonuclease function of T4 DNA polymerase compared to non-treated oligonucleotides, suggesting structural distortions of DNA caused by acetaldehyde-treatment. Neither in vitro DNA synthesis reactions of phi29 DNA polymerase nor in vitro RNA synthesis reactions of T7 RNA polymerase were observed when acetaldehyde-treated plasmid DNA was used, compared to when non-treated plasmid DNA was used, suggesting that acetaldehyde-induced DNA lesions inhibited replication and transcription in DNA metabolism. Conclusions Acetaldehyde-induced DNA lesions could affect the relative resistance to endo- and exo-nucleolytic activity and also inhibit in vitro replication and in vitro transcription. Thus, investigating the effects of acetaldehyde-induced DNA lesions may enable a better understanding of the toxicity and carcinogenicity of acetaldehyde.

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