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RNA
Published in Paul Pumpens, Single-Stranded RNA Phages, 2020
A large, complex study was performed on the addressed fragmentation of the MS2 RNA by efficient collaboration of laboratories from Moscow and Riga (Metelev et al. 1978, 1980a,b; Stepanova et al. 1979). In this study, ribonuclease H from E. coli was used for selective cleavage of MS2 and R17 RNAs in the region of a heteroduplex formed in RNA with an oligodeoxyribonucleotide complementary to a certain part of the RNA. It was shown that ribonuclease H split the molecule of RNA in the position corresponding to the 3′-end of the heteroduplex. Later, this approach was developed further by crosslinking of 15-mer DNA fragments to the Thermus thermophilus ribonuclease HI at different enzyme positions (Chon et al. 2002). As a result, site-specific cleavage of the MS2 RNA was achieved successfully by this thermostable DNA-linked ribonuclease H.
Cellular and Molecular Basis of Human Biology
Published in Lawrence S. Chan, William C. Tang, Engineering-Medicine, 2019
DNA PCR. To perform DNA PCR, researchers need to prepare several essential components: The DNA target template, a designed primer pair that is complementary to DNA target at each end of desired DNA segment to be amplified, heat-stable DNA polymerase commonly Taq polymerase that is derived from a hot-spring-grown bacteria Thermus aquatica, mixture of deoxynucleoside triphosphate, and a bivalent cation-containing buffer solution. Magnesium is the common cation used. Of note, the bacteria Thermus aquatica was found, isolated, and named by an American Scientist Dr. Thomas D. Brock and his student when they studied microorganisms at a large hot spring near the Great Fountain Geyser inside the Yellowstone National Park (Brock 1994).
Molecular biology
Published in Maxine Lintern, Laboratory Skills for Science and Medicine, 2018
The basic principle of the method exploits the fact that DNA is double-stranded, and if you denature the helix using high temperatures, you can make another complementary strand using DNA polymerase and a few loose bases, effectively doubling the amount of DNA you have. If you do that again and again, you can see how you can make lots of copies, all identical to the original strand. What makes this technique practical is the use of automation by engaging the services of the highly temperature-resistant DNA polymerase from Thermus aquaticus. This bacteria lives in hot springs, and is quite happy at temperatures around 100°C. Most other DNA polymerases only work at low temperatures, but DNA only denatures into two strands at high temperatures. Thus Taq polymerase can be used to make copies of DNA by cycling between high and low temperatures every few minutes, so in the space of a few hours you can amplify one copy of DNA up to millions! PCR machines costing a few thousand pounds are fundamentally programmable heating/cooling blocks that take your samples up and down to exact temperatures for an exact number of cycles, before cooling the amplified DNA ready for use. Many labs with high usage of PCR techniques will have banks of machines running 24 hours a day.
Modeling spatial interaction networks of the gut microbiota
Published in Gut Microbes, 2022
Xiaocang Cao, Ang Dong, Guangbo Kang, Xiaoli Wang, Liyun Duan, Huixing Hou, Tianming Zhao, Shuang Wu, Xinjuan Liu, He Huang, Rongling Wu
Only three positions, transverse colon, ileum, and cecum, were sampled for the HC group, and these positions display distinct network topologies (Figure 4). Each of these three HC networks is tremendously different from the UC network at the same position. In general, the HC networks are well mixed by commensalism and amensalism, whereas the UC networks tend to be dominated by commensalism, suggesting that a healthy network can better balance different types of microbes than a diseased network. At the cecum and ileum positions, Firmicutes establishes a commensalistic relationship toward Proteobacteria in the UC networks, but an amensalistic relationship between these two phyla is detected in the HC networks. In the transverse colon network, although commensalism occurs from Firmicutes to Proteobacteria in both UC and HC groups, the strength of this interaction is larger in the former than in the latter. Also, Deinococcus-Thermus is commensalistic toward Fusobacteria in the UC networks, but this relationship is amensalistic in the HC networks. Taken together, the strength, pattern and architecture of microbial interactions vary spatially across the biogeographic positions of the gut, with the degree of variation depending on healthy state. Network analysis on the three commonly measured positions suggests that healthy networks display a greater position-dependent variability than diseased networks. A series of spatially reconstructed networks can more precisely characterize key microbial interactions that interrogate why and how a healthy state becomes unhealthy.
Advances in biocatalytic and chemoenzymatic synthesis of nucleoside analogues
Published in Expert Opinion on Drug Discovery, 2022
Sebastian C. Cosgrove, Gavin J. Miller
Relatedly, a PyNP from Thermus thermophilus displayed a broad activity profile (pH 4–10, operational temperatures of up to 100 °C), but its application was limited by nucleobase inhibition at low concentrations [19]. Transglycosylation has also been used recently by the same group to effect the synthesis of 2-selenopyrimidines [20], the inclusion of which within wider nucleic acid chemistries has an established capability to deliver structure-to-function tools [21]. Enzymatic synthesis of 2-selenouridine and 2-selenothymidine was effected using uridine and thymidine as donors and a 2-selenyl heterobase. Significantly larger equilibrium constants were observed for these non-native transglycosylation processes, requiring an excess of the nucleoside donor to access reaction yields of 40–48%.
Molecular regulation of adhesion and biofilm formation in high and low biofilm producers of Bacillus licheniformis using RNA-Seq
Published in Biofouling, 2019
Faizan Ahmed Sadiq, Steve Flint, Hafiz Arbab Sakandar, GuoQing He
Sugars and fatty acids in the biofilm matrix of B. licheniformis appear to play an important role in the biofilm matrix. GalE plays a major role in biofilm formation by epimerising UDP-N-acetylgalactosamine and UDP-galactose for the biosynthesis of EPS. It has previously been demonstrated that GalE plays a very crucial role in biofilm formation by Thermus thermophilus (Niou et al. 2009), V. cholerae (Nesper et al. 2001) and B. subtilis (Chai et al. 2012). Many bacterial species like Neisseria meningitides and Neisseria gonorrhoeae do not use UDP-galactose as a carbon source, but use it only for the synthesis of EPS (Jennings et al. 1993; Robertson et al. 1993). The overexpression of four genes encoding UDP-glucose 4-epimerase GalE in the biofilm state of H strain compared to only one gene in the L strain highlights its role in EPS production in B. licheniformis (Table 2).