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Anhydrotetracycline

Anhydrotetracycline is a derivative of the antibiotic tetracycline.
It is used as a research tool to study the structure and function of tetracycline-binding proteins.
Anhydrotetracycline lacks the hydroxyl group present in tetracycline, which can affect its binding affinity and biological activity.
Researchers utilize anhydrotetracycline to investigate the molecular mechanisms of tetracycline action and resistance, contributing to a better understanding of this important class of antibiotics.
The PubCompare.ai platform can help identify the most accurate and reproducible protocols for working with anhydrotetracycline, optimizing your research for accuracy and reprodcibility.

Most cited protocols related to «Anhydrotetracycline»

1
Cultivation of Thalassiosira pseudonana in Artificial Seawater
Cited 44 times
Parasites were grown in hTERT-BJ1 (clontech) cells in supplemented Dulbecco's modified Eagle's medium [67] ). Parasite cloning and plaque assays were performed in human foreskin fibroblasts (HFF). For the selection of stable transgenic lines, drugs were added as follow: 1 µM pyrimethamine added one day after transfection for one week, 20 µM chloramphenicol added the day of transfection for three weeks, 5 µM FUDR added two days after transfection for one week. To repress the regulated promoter, parasites were grown in the presence of 0.5 µM anhydrotetracycline (ATc).
Thalassiosira pseudonana (Hustedt) Hasle et Heimdal CCMP1335 was grown in an artificial seawater medium (EASW) according to the North East Pacific Culture Collection protocol (http://www3.botany.ubc.ca/cccm/NEPCC/esaw.html) at 18°C under constant light. Where indicated, NaNO3 was omitted from the medium (nitrogen-free medium) or replaced by 0.55 mM NH4Cl (ammonium medium).
Sheiner L., Demerly J.L., Poulsen N., Beatty W.L., Lucas O., Behnke M.S., White M.W, & Striepen B. (2011). A Systematic Screen to Discover and Analyze Apicoplast Proteins Identifies a Conserved and Essential Protein Import Factor. PLoS Pathogens, 7(12), e1002392.
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Publication 2011
Ammonium anhydrotetracycline Animals, Transgenic Biological Assay Cells Chloramphenicol Fibroblasts Floxuridine Foreskin Homo sapiens Light Nitrogen Parasites Pharmaceutical Preparations Pyrimethamine Senile Plaques Transfection
2
Mycobacterium tuberculosis Strain H37Rv
Cited 34 times
MTB H37Rv was used for all experiments with the single exception of one experiment performed in M. smegmatis (Supplementary Fig. 21). This MTB strain was fully sequenced by the Broad Institute (GI:397671778). For Chip-Seq, cells were cultured in Middle brook 7H9 with ADC (Difco), 0.05% Tween 80, and 50 µg ml−1 hygromycin B at 37 °C with constant agitation and induced with 100 ng ml−1 anhydrotetracycline (ATc) during mid-log-phase growth, and ChIP was performed using a protocol optimized for mycobacteria and related Actinomycetes. For the hypoxia and re-aeration time-course, bacilli were cultured in bacteriostatic oxygen-limited conditions (1% aerobic O2 tension) for seven days, followed by re-aeration. Bacteria were cultured in Sauton’s medium without detergent or exogenous lipid source. Profiling samples were collected as described in the Supplementary Text. All data available at http://TBDB.org. Expression data also available at GEO (accession number GSE43466).
Galagan J.E., Minch K., Peterson M., Lyubetskaya A., Azizi E., Sweet L., Gomes A., Rustad T., Dolganov G., Glotova I., Abeel T., Mahwinney C., Kennedy A.D., Allard R., Brabant W., Krueger A., Jaini S., Honda B., Yu W.H., Hickey M.J., Zucker J., Garay C., Weiner B., Sisk P., Stolte C., Winkler J.K., Van de Peer Y., Iazzetti P., Camacho D., Dreyfuss J., Liu Y., Dorhoi A., Mollenkopf H.J., Drogaris P., Lamontagne J., Zhou Y., Piquenot J., Park S.T., Raman S., Kaufmann S.H., Mohney R.P., Chelsky D., Moody D.B., Sherman D.R, & Schoolnik G.K. (2013). The Mycobacterium tuberculosis regulatory network and hypoxia. Nature, 499(7457), 178-183.
Publication 2013
Actinomycetes anhydrotetracycline Bacteria Bacteria, Aerobic Cells Chromatin Immunoprecipitation Sequencing Culture Media Detergents DNA Chips Hygromycin B Hypoxia Lacticaseibacillus casei Lipids Mycobacterium Strains Tween 80
3
Deletion Constructs and Complementation for Staphylococcus Genes
Cited 29 times
Deletion constructs for the hsdR, sauUSI, sauUSIECORV, and mcrR genes and a putative nudix hydrolase-encoding gene (Table 1) were PCR amplified as follows. A sequence upstream of the gene to be deleted was amplified with oligonucleotides A and B (A/B) (up to the start codon) and the downstream sequence with oligonucleotides C/D (down from the stop codon) separately. The upstream and downstream PCR products were diluted 1:20, and 1 µl of each was used as the template in a second SOE PCR with the A/D primers. Deletion constructs were cleaved at endonuclease sites introduced into A and D primers during PCR and ligated into pIMAY cut with the same enzymes and then transformed into E. coli DC10B. The plasmid DNA was sequenced. The DNA was then electroporated into the target strain and plated onto BHIA plus Cm10 at 28°C.
To correct the premature stop codon in the sauUSI gene of S. aureus RN4220, the wild-type sequence from S. aureus 8325-4 was amplified as a 1-kb fragment centered on the RN4220 premature stop codon and then processed as described above.
To complement the sauUSI mutation in S. aureus NRS384 hsdRINT sauUSIINT, the sauUSI deletion mutant was reverted to the wild type by allelic exchange. To differentiate NRS384 hsdRINT from the NRS384 hsdRINT sauUSIECORV complemented strain, a new EcoRV restriction site (http://emboss.bioinformatics.nl/cgi-bin/emboss/silent) was introduced into the complementation construct without altering the coding sequence. Phenotypically, no differences were observed between NRS384 hsdRINT and NRS384 hsdRINT sauUSIECORV mutants.
To integrate pIMAY into the chromosome, a single colony from the transformation plate was homogenized in 200 µl of TSB. The suspension was diluted 10-fold to 10−3, and 100 µl of each dilution was spread on BHIA plus Cm10 and incubated overnight at 37°C. For S. epidermidis, a colony from the transformation plate was inoculated into BHI plus Cm10 and grown overnight at 37°C, and diluted suspensions of the bacterial growth were plated for single colonies. For both S. aureus and S. epidermidis, large colonies were streaked on BHIA plus Cm10 and incubated overnight at 37°C, and colony PCR analysis was performed to determine (i) the absence of extrachromosomal plasmid DNA (with MCS oligonucleotides IM151/152) (Fig. 3B) and (ii) whether plasmid integration had occurred upstream or downstream of the gene (OUT F/D Rev oligonucleotides or OUT R/A Fwd oligonucleotides, e.g., hsdR IM5/94 or IM6/93) (Fig. 3C). Overnight cultures of both the upstream or downstream crossover that were free of replicating plasmid were grown at 28°C without chloramphenicol and then plated onto BHIA containing 1 µg/ml anhydrotetracycline (Vetranal; Sigma) (BHIA plus ATc). The plates were incubated at 28°C for 2 days. Large colonies were patched on BHIA plus ATc and BHIA plus Cm10 and grown at 37°C overnight. Chloramphenicol-sensitive colonies were screened by colony PCR with oligonucleotides to identify clones containing the desired mutation (OUT F/OUT R [e.g., ∆hsdR-IM5/6]). Putative mutants were validated by PCR amplification of genomic DNA flanking the deletion and DNA sequencing.
Monk I.R., Shah I.M., Xu M., Tan M.W, & Foster T.J. (2012). Transforming the Untransformable: Application of Direct Transformation To Manipulate Genetically Staphylococcus aureus and Staphylococcus epidermidis. mBio, 3(2), e00277-11.
Publication 2012
4
Mycobacterial Growth and Genetic Manipulation
Cited 25 times
Table 1 lists the strains and plasmids used in this study. Mycobacteria were grown in Middlebrook 7H9 medium (Difco/VWR) with 0.2% glycerol and 0.05% Tween-80. For growth of M.tuberculosis H37Rv, the medium was supplemented with 0.5% BSA, 0.2% dextrose and 0.085% sodium chloride (ADN). For selection of recombinant mycobacteria, kanamycin and hygromycin were used at 30 and 50 μg/ml, respectively. Escherichia coli DH5α was grown in Luria–Bertani broth (LB) and kanamycin and hygromycin were used at 60 and 200 μg/ml, respectively.
Ehrt S., Guo X.V., Hickey C.M., Ryou M., Monteleone M., Riley L.W, & Schnappinger D. (2005). Controlling gene expression in mycobacteria with anhydrotetracycline and Tet repressor. Nucleic Acids Research, 33(2), e21.
Publication 2005
Escherichia coli Glucose Glycerin hygromycin A Kanamycin Mycobacterium Mycobacterium tuberculosis H37Rv Plasmids Sodium Chloride Strains Tween 80
5
Tet-Regulated Gene Expression in Mycobacteria
Cited 24 times
Tet operators were inserted into Psmyc using oligonucleotide-directed PCR mutagenesis (20 ). The PCR oligonucleotides were smyc-tet1 (5′-gagtttgtcctccctatcagtgatagataggctctgggagtacccgtctg-3′), smyc-tet2 (5′-ctgatagggaggacaaactctatcactgatagggagttctcccgctcgtcagagaccct-3′), gfp2 (5′-gcatcaccttcaccctctccact gac-3′) and poly3 (5′-gaactagttgattagctaagcagaagg-3′). A PCR product containing Pmyc1tetO, the tetO containing Psmyc derivative, was digested with XbaI and SphI and cloned into pUV15 to replace Psmyc. A Tn10-derived tetR gene was amplified by PCR from pWH520 (27 (link)) and cloned into pMS2 for extrachromosomal expression or into pMV306Km for integration onto the mycobacterial chromosome. The promoters Psmyc or Pimyc were used for the expression of TetR in mycobacteria. All constructs were verified by restriction analysis and DNA sequencing.
Ehrt S., Guo X.V., Hickey C.M., Ryou M., Monteleone M., Riley L.W, & Schnappinger D. (2005). Controlling gene expression in mycobacteria with anhydrotetracycline and Tet repressor. Nucleic Acids Research, 33(2), e21.
Publication 2005
Chromosomes Genes Mycobacterium Oligonucleotide-Directed Mutagenesis Oligonucleotides PMS2 protein, human

Most recents protocols related to «Anhydrotetracycline»

1
Tet(56-2) and Tet(56-3) Inhibitor Potency Assay
All experiments were prepared open to air in non-degassed buffer solutions at room temperature. Half-maximal inhibitory concentrations (IC50) for the inhibition of Tet(56-2) and Tet(56-3) were determined from the velocities of substrate degradation in the presence of varying concentrations of inhibitor. Reaction samples were prepared in 100 mM TAPS buffer (pH 8.5) with 504 μM NADPH, 5.04 mM MgCl2, 25.3 μM substrate, varying concentrations of inhibitor (0-227 μΜ), and 0.4 μM enzyme (final working concentrations). Reactions were initiated by the addition of enzyme and were monitored continuously via optical absorbance spectroscopy at 400 nm for 2 min (performed in triplicate as independent trials). Initial enzyme velocities were determined by linear regression using Agilent Cary WinUV Software over the linear range of the reaction (0 to 1 min). The velocities were plotted against the logarithm of inhibitor concentration, and apparent IC50 values were determined using nonlinear regression analysis in GraphPad Prism 6. Each set of experiments included a no-TDase control reaction which was used as the full enzyme inhibition velocity and assigned to inhibitor concentration of 1 × 1015, and a no-inhibitor control which was assigned an inhibitor concentration of 1 × 10–15. A no-tetracycline control was also performed to search for potentially competitive background signals generated from the enzymatic degradation of the inhibitor. For all inhibitor-enzyme combinations, the initial velocities of the no-tetracycline controls were negligible.
Blake K.S., Kumar H., Loganathan A., Williford E.E., Diorio-Toth L., Xue Y.P., Tang W.K., Campbell T.P., Chong D.D., Angtuaco S., Wencewicz T.A., Tolia N.H, & Dantas G. (2024). Sequence-structure-function characterization of the emerging tetracycline destructase family of antibiotic resistance enzymes. Communications Biology, 7, 336.
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Publication 2024
2
Investigating Temperature and Nutrient Effects on SCR
Plasmids encoding the reporters for SCR events were electrotransformed in the wild-type K-12 MG1655 E. coli strain. Transformants were grown on LB-agar plates and inoculated into 384-well plates containing LB media supplemented with 15 µg/mL of chloramphenicol. To store the transformants at −80 °C (glycerol stock), they were grown at 37 °C to saturation, and glycerol was added until a final concentration of 20%.
To investigate the effect of temperature on SCR, E. coli cultures were grown into 384-well plates without shaking under saturated humidity conditions at 18, 25, 37, and 42 °C to saturation. To address the nutrient depletion effect on SCR, LB and M9 media were tested.
To titrate the expression of the Gly95-TGA reporter, 0, 25, 50, 100, 200, 400, and 800 µg/L of anhydrotetracycline was added to the media. For the library study, to induce the expression, 400 µg/L of anhydrotetracycline was added to the media. Cells were grown under light protection to avoid the photodegradation of the anhydrotetracycline.
Romero Romero M.L., Poehls J., Kirilenko A., Richter D., Jumel T., Shevchenko A, & Toth-Petroczy A. (2024). Environment modulates protein heterogeneity through transcriptional and translational stop codon readthrough. Nature Communications, 15, 4446.
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Publication 2024
3
Checkerboard Whole Cell Inhibition Assay
For checker-board whole cell inhibition assays, anhydrotetracycline was two-fold serially diluted in a constant concentration of tetracycline. Liquid cultures of each strain were grown to exponential phase then diluted to a standard concentration (OD600 = 0.0015, which is equivalent to double ~5 × 105 CFU/mL) and inoculated into each panel at a 1:1 ratio. Thus, each well had a final concentration of 50 μg/mL kanamycin, 1 mM IPTG, ~5 × 105 CFU/mL (0.5 MacFarland) cells, and variable concentrations of the antibiotic of interest or anhydrotetracycline. Each strain-antibiotic/inhibitor combination was tested in triplicate, along with no-drug and no-cell controls. Inoculated panels were sealed with Breathe-Easy membranes (Sigma-Aldrich) and incubated at 37 °C for 20 h. Panels were scored by absorbance measurements at 600 nm (OD600) using the Synergy H1 microplate reader (Biotek Instruments, Inc).
Blake K.S., Kumar H., Loganathan A., Williford E.E., Diorio-Toth L., Xue Y.P., Tang W.K., Campbell T.P., Chong D.D., Angtuaco S., Wencewicz T.A., Tolia N.H, & Dantas G. (2024). Sequence-structure-function characterization of the emerging tetracycline destructase family of antibiotic resistance enzymes. Communications Biology, 7, 336.
Full text: Click here
Publication 2024
4
Purification of C-terminally Tagged SasG
The sasG gene from S. aureus MW2 was amplified using primers HC416 and HC418 (Table 2), which remove the last 33 amino acids of SasG, including the LPXTG cell wall anchor, and replace them with a glycine followed by six histidine residues. This C-terminally tagged, secreted version of sasG was cloned into pALC2073 under the control of an anhydrotetracycline-inducible promoter, generating pHC90. We decided to purify this version of SasG from S. aureus LAC, which does not have an intact copy of sasG on the chromosome. To avoid potential proteolysis, we used a previously developed strain of LAC lacking secreted proteases (AH1919). In addition, we modified AH1919 to be resistant to anhydrotetracycline by integrating the empty vector pLL29 (95 (link)) in the phage 11 attachment site, generating host strain AH4607.
For expression of SasG, pHC90 was moved into AH4607 and a 5 mL culture was grown overnight at 37°C in TSB with chloramphenicol. This overnight culture was used to inoculate 1 L of TSB supplemented with chloramphenicol and 0.15 µg/mL anhydrotetracycline. The culture was grown with shaking for ~6.5 h at 37°C. Cells were removed by centrifugation, and the culture supernatant was concentrated to ~30 mL using an Amicon stirring pressure concentrator with a 100 kDa cutoff filter. The supernatant was dialyzed twice against binding buffer (50 mM sodium phosphate, 300 mM NaCl, pH 8). SasG-His6 was then purified using a pre-packed 5 mL IMAC cartridge (Bio-Rad) on a Bio-Rad FPLC. SasG-His6 was eluted with a linear gradient up to 100% elute buffer (50 mM sodium phosphate, 300 mM NaCl, 250 mM imidazole, pH 8). The protein was then concentrated and dialyzed against the storage buffer (20 mM sodium phosphate, 150 mM NaCl, pH 7.5). Glycerol was added to 20% before flash freezing and storing at −80°C.
Crosby H.A., Keim K., Kwiecinski J.M., Langouët-Astrié C.J., Oshima K., LaRivière W.B., Schmidt E.P, & Horswill A.R. (2024). Host-derived protease promotes aggregation of Staphylococcus aureus by cleaving the surface protein SasG. mBio, 15(4), e03483-23.
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Publication 2024
5
Comprehensive Tetracycline Quality Analysis
Tetracycline HCl USP RS (Sigma, China) and USP RS standard impurities (Epitetracycline HCl (USA), Anhydrotetracycline HCl (India), and 4-Epi-anhydrotetracycline HCl (India)) were used in this work. All USP RS standards were kindly donated by the United States Pharmacopoeia, Maryland, USA. Tetracycline HCl BP working standard (98.24% potency) was kindly supplied by National Veterinary Institute (NVI) of Ethiopia. Three brands of tetracycline HCl 1% eye ointment preparations (Tetracycline HCl USP 1% eye ointment (Shanghai General Pharmaceutical Co Ltd, China), Galentic 1% eye ointment (Galentic Pharma Pvt. Ltd, India), Brassica 1% eye ointment (Brassica Pharma Pvt. Ltd, India)) and two brands of tetracycline HCl 3% skin ointment preparations (Galantic 3% skin oint (Galentic PharmaPvt.Ltd, India), Aurocycline 3% skin ointment (Aurochem Laboratories Pvt. Ltd, India)) and tetracycline HCl API (China), donated by NVI, Ethiopia, were used. Eye and skin ointments were purchased from pharmacy retail outlets in Addis Ababa, the capital city of Ethiopia. Further detailed information about standards and dosage forms has been given in the Additional file 1: Table S1.
Gashaw M., Layloff T., Hymete A, & Ashenef A. (2024). Stability indicating high performance thin layer chromatography method development and validation for quantitative determination of tetracycline hydrochloride in tetracycline hydrochloride active pharmaceutical ingredient (API) and its dosage forms. BMC Chemistry, 18(1), 82.
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Publication 2024

Top products related to «Anhydrotetracycline»

1
Anhydrotetracycline by Merck Group
Sourced in United States, Germany
Anhydrotetracycline is a chemical compound used as a laboratory reagent. It is a derivative of the tetracycline antibiotic family. Anhydrotetracycline is primarily utilized in research and scientific applications, rather than for direct medical or clinical use.
2
Anhydrotetracycline atc by Merck Group
Sourced in United States
Anhydrotetracycline (ATc) is a synthetic derivative of the tetracycline class of antibiotics. It is used as a lab equipment product for the regulation of gene expression in biological systems.
3
Anhydrotetracycline atc by Takara Bio
Sourced in United States
Anhydrotetracycline (aTc) is a chemical compound used as a tool in molecular biology and biotechnology. It serves as a synthetic inducer, providing a means to regulate gene expression in genetically engineered organisms.
4
Kanamycin by Merck Group
Sourced in United States, Germany, United Kingdom, France, Spain, China, Israel, India, Canada, Macao, Australia, Sao Tome and Principe, Belgium, Sweden, Poland, Japan, Switzerland, Brazil, Italy, Ireland
Kanamycin is a broad-spectrum antibiotic derived from the bacterium Streptomyces kanamyceticus. It is commonly used as a selective agent in molecular biology and microbiology laboratories for the growth and selection of bacteria that have been genetically modified to express a gene of interest.
5
Anhydrotetracycline by Cayman Chemical
Sourced in United States
Anhydrotetracycline is a chemical compound that is commonly used in laboratory settings. It is a derivative of the tetracycline class of antibiotics and is often utilized for research and analytical purposes.
6
Kanamycin by GoldBio
Sourced in United States
Kanamycin is an aminoglycoside antibiotic used as a selective agent in various molecular biology techniques. It functions by inhibiting protein synthesis in bacterial cells.
7
Anhydrotetracycline hydrochloride atc by Merck Group
Sourced in Macao
Anhydrotetracycline hydrochloride (aTc) is a chemical compound used as a laboratory reagent. It functions as a tetracycline derivative and can be used for various applications in scientific research and experiments.
8
Chloramphenicol by Merck Group
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Chloramphenicol is a bacteriostatic antibiotic that inhibits protein synthesis in bacteria. It is commonly used in microbiology laboratories for selective cultivation and identification of bacterial species.
9
L arabinose by Merck Group
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L-arabinose is a monosaccharide that serves as a common laboratory reagent. It is a colorless crystalline solid that is soluble in water and has the molecular formula C₅H₁₀O₅.
10
Arabinose by Merck Group
Sourced in United States, Germany, China, Sao Tome and Principe, United Kingdom, Sweden
Arabinose is a monosaccharide that is commonly used as a component in various laboratory equipment and supplies. It functions as a carbohydrate source and can be utilized in various biochemical and microbiological applications.

More about "Anhydrotetracycline"

Anhydrotetracycline, also known as aTc or ATc, is a derivative of the antibiotic tetracycline that lacks the hydroxyl group present in its parent compound.
This structural modification can affect the binding affinity and biological activity of anhydrotetracycline compared to tetracycline.
Researchers utilize anhydrotetracycline as a valuable tool to study the structure and function of tetracycline-binding proteins, contributing to a better understanding of the molecular mechanisms of tetracycline action and resistance.
Anhydrotetracycline is often used in conjunction with other compounds, such as kanamycin, chloramphenicol, and L-arabinose, to investigate various aspects of bacterial physiology and gene regulation.
For example, the arabinose-inducible promoter system, which employs L-arabinose as an inducer, can be used in combination with anhydrotetracycline to control gene expression in a tightly regulated manner.
The PubCompare.ai platform can help researchers identify the most accurate and reproducible protocols for working with anhydrotetracycline, optimizing their research for accuracy and rpodocibility.
By leveraging the power of AI-driven protocol optimization, scientists can access the best products and procedures, ensuring their experiments yield reliable and meaningful results.