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Mycobacterium tuberculosis H37Rv

Mycobacterium tuberculosis H37Rv is a widely used laboratory strain of the bacterium that causes tuberculosis.
This virulent strain is commonly employed in research to study the pathogenesis, immunology, and drug development for tuberculosis.
PubCompare.ai can optimize your M. tuberculosis H37Rv research by helping you identify the most effective protocols from published literature, preprints, and patents.
With side-by-side comparisons, you can streamline your studies and enhance reproducibility.
Leverage the power of AI to improve the accuracy and effciency of your M. tuberculosis H37Rv research.

Most cited protocols related to «Mycobacterium tuberculosis H37Rv»

Rapid Annotation Transfer Tool (RATT)

RATT is programmed in ‘bash’ and ‘PERL’ and its design is illustrated in Figure 1 and Supplementary Figure S1. First, two sequences are compared using ‘nucmer’ from the MUMmer package (17 (link)) to define sequence regions that share synteny. Those regions are filtered using configurable parameters depending on the type of annotation mapping that is being attempted. Preset parameters are provided for transfers between assembly versions, strains or species (see Supplementary Table S1). To be included, the minimum nucleotide sequence identity between synteny blocks must be 40%. Synteny information is stored as a base range in the query and its associated base range in the reference. However, this information alone is inadequate to map the annotation because insertions or deletions (indels) change the relative distance between mapped synteny blocks. The coordinates are therefore sequentially adjusted across a synteny block by calling indels using ‘show-snp’ from the MUMmer package. Accurately calling indels within repetitive regions presents a particular challenge. Therefore, RATT recalibrates the adjusted coordinates using single nucleotide polymorphisms (SNPs, also called using ‘show-snp’) as unambiguous anchor points within synteny blocks. In transfers between very closely related sequences (e.g. successive assembly versions), SNPs may occur with insufficient frequency to perform this coordinate adjustment. In such cases, RATT modifies the query by inserting a faux SNP every 300 bp to aid in the recalibrating step. The final sequence and transferred annotations remain unaffected.
Figure 1.

Workflow of RATT.

Once the coordinates within synteny blocks have been defined, RATT proceeds to the annotation-mapping step, whereby each feature within a reference EMBL file is associated with new coordinates on the query (Supplementary Figure S1B). A feature is not mapped (and is put in the non-transferred bin file), if it bridges a synteny break and if its coordinate boundaries match different chromosomes, different DNA strands, or if the new mapped distance of its coordinates has increased by more than 20 kb. If a short sequence from the beginning, middle or the end of a feature can be placed within a synteny region, mapping is attempted (see Supplementary Figure S1B). In addition, if the exons of a single gene model map to different gene regions, the model is split and identified in the output file. The bin is an EMBL-format file that can be loaded onto the reference sequence for analysis (see Figure 2, brown colour track). Further outputs include statistics about transferred features or the amount of synteny conserved between the reference and query, as well as Artemis-readable files showing SNPs, indels and regions that lack synteny between the compared sequences, see the example on the sourceforge site.
Figure 2.

Transfer of annotation from the M. tuberculosis strain H37Rv onto the strain F11 sequence, over a deletion. The genomes of H37Rv (upper) and F11 (lower) are shown using the Artemis Comparison Tool (ACT). The source H37Rv annotation (light blue) is directly mapped onto F11 by RATT (green) except for those features corresponding to a region that is unique to the source strain that cannot be transferred and are written to a separate output file (brown).

Although two sequences may be related, differences can occur, such as a change in the start or stop codons of a protein-coding sequence. Therefore, we implemented a correction algorithm in RATT (see Supplementary Figure S1C). Figure 3 shows examples of the correction step. First, the start codon is checked. If it is not present, the upstream sequence is searched for a new start codon (Figure 3A). If a stop codon is found, the first start codon downstream is used. In the absence of any start codon, an error is recorded in the results file. If the sequence between exons has no stop codon and a length divisible by three bases but the splice acceptor or donor sequences are wrong, then the intron is eliminated. Likewise, frameshifts previously introduced into the reference to maintain conceptual translations (for instance, in apparent pseudogenes) will also be removed from coding sequences in the query. RATT will also detect, and attempt to fix, incorrect splice sites. As splice sites are difficult to annotate correctly, RATT only tries to correct a gene model that has one wrong splice site. If one incorrect splice site is detected, the closest alternative splice donor or acceptor is found that, when used, generates no frame shifts. Next, RATT searches for genes or exons with internal stop codons, further than 150 bp from the 3′-end. If the introduction of a frameshift would generate a model without internal stop codons, the model is corrected (Figure 3C). Stop codons are corrected last: if a model has less than five internal stops in its last exon, the model is shortened to the first stop codon (Figure 3B). If the model has no stop codon it is extended downstream until a stop codon is found.
Figure 3.

RATT corrections of transferred annotations. Annotation from H37Rv were transferred onto the F11 sequence (pale blue), corrected (green) and then compared with the existing strain F11 annotation in EMBL (yellow). (A and B) The correction of start and stop codons, respectively. In a more complex mapping situation (C), where all three reading frames are shown for clarity, RATT maps a large single coding sequence (CDS) from H37Rv to a locus within F11 that includes several in-frame stop codons. By inserting a frameshift (i.e. to indicate a pseudogene) the conceptual translation is preserved. This contrasts with two overlapping genes predicted as part of the F11 genome project.

Different criteria can be specified depending on the translation that an organism uses (e.g. such bacterial TTG and GTG start codons) or whether unsual splice sites are used. RATT is programmed in PERL and was tested in UNIX/LINUX environments. The output can be loaded into Artemis/Act. The list and explanation of all the output files can be found at the sourceforge site.

Otto T.D., Dillon G.P., Degrave W.S, & Berriman M. (2011). RATT: Rapid Annotation Transfer Tool. Nucleic Acids Research, 39(9), e57.

Publication 2011

Bacteria Base Sequence Chromosomes Codon, Initiator Codon, Terminator Contrast Media Deletion Mutation Exons Frameshift Mutation Gene Deletion Genes Genes, Overlapping Genome INDEL Mutation Insertion Mutation Introns Light Microtubule-Associated Proteins Mycobacterium tuberculosis H37Rv Open Reading Frames Pseudogenes Reading Frames Repetitive Region Single Nucleotide Polymorphism Strains Synteny Tissue Donors

Evaluating RATT genome annotation transfer

To evaluate RATT, we assessed its performance using manually annotated genomes. The M. tuberculosis strain H37Rv (NCBI:AL123456) was used to annotate the genome of strain F11 using the ‘Strain’ comparison option. Results were compared with the existing annotation of F11 (NCBI:CP000717). In addition, the annotation of P. chabaudi was mapped to the P. berghei version 9 genome using the ‘Species’ comparison option. The P. chabaudi/P. berghei dataset can be downloaded from http://ratt.sourceforge.net/Chab_berg.zip. The files relating to the transfer of annotation between P. berghei assemblies can be found at http://ratt.sourceforge.net/Berg_berg.zip. The transfer was performed using the ‘Assembly.Repetitive’ parameters, and the results are included in the latest GeneDB version (http://www.genedb.org).
Although, direct benchmarking was not possible because RATT presents a new strategy, we ran Glimmer3—a popular ab initio gene predictor—on the tuberculosis dataset as a comparator. Particular attention was given to the number of CDSs transferred or predicted, and whether their boundaries coincided with curated models. After running RATT on each of the three datasets, the transferred annotations were manually checked in Artemis and ACT.

Otto T.D., Dillon G.P., Degrave W.S, & Berriman M. (2011). RATT: Rapid Annotation Transfer Tool. Nucleic Acids Research, 39(9), e57.

Publication 2011

Attention Genes, vif Genome Mycobacterium tuberculosis H37Rv Strains Triglyceride Storage Disease with Ichthyosis Tuberculosis

Mycobacterial and E. coli Culture Conditions

M. smegmatis mc²155 [72] (link), M. tuberculosis H37Rv and Escherichia coli NEB-10β (New England Biolabs UK Ltd) were used in this work. M. smegmatis and M. tuberculosis were grown on Middlebrook 7H11 agar medium (BD Diagnostics) supplemented with 0.5% glycerol and 10% oleic acid albumin-dextrose-catalase (OADC) (BD Diagnostics). When required, filter-sterilised luciferin was added at a final concentration of 0.157 mM. Liquid cultures of M. smegmatis and M. tuberculosis were grown either in Middlebrook 7H9 broth (BD Diagnostics) containing 0.05% Tween 80 (Sigma) and 10% albumin-dextrose-catalase (ADC) enrichment (BD Diagnostics), or (for M. smegmatis Gluc assays) in Luria-Bertani (LB) medium with 0.05% Tween. LB medium was preferred for the Gluc assays because the background of coelenterazine was 100 times lower in that medium than in 7H9 broth. LB medium was used for culturing E. coli. All the strains were grown at 37°C. The following antibiotics were added when appropriate: ampicillin [100 µg ml⁻¹ (Sigma)], hygromycin B [150 µg ml⁻¹ (Invitrogen)] and kanamycin [25 µg ml⁻¹, for mycobacteria, 50 µg ml⁻¹ for E. coli (Sigma)].

Andreu N., Zelmer A., Fletcher T., Elkington P.T., Ward T.H., Ripoll J., Parish T., Bancroft G.J., Schaible U., Robertson B.D, & Wiles S. (2010). Optimisation of Bioluminescent Reporters for Use with Mycobacteria. PLoS ONE, 5(5), e10777.

Publication 2010

Agar Albumins Ampicillin Antibiotics, Antitubercular Biological Assay Catalase coelenterazine Diagnosis Escherichia coli Glucose Glycerin Hygromycin B Kanamycin Luciferins Mycobacterium Mycobacterium tuberculosis Mycobacterium tuberculosis H37Rv Oleic Acid Strains Tween 80 Tweens

Construction and Characterization of Mycobacterial Reporter Strains

The plasmids used in this study are described in Tables 1–4. The integrating expression vectors pMV306hsp and pMV306myc were constructed by cloning into pMV306 the promoters P_hsp60 and P_myc1tetO obtained as NotI-HindIII and XbaI-SalI fragments from pSMT3 and pSE100 respectively. All reporter genes, except for the lux operon, were PCR amplified using the primers and templates listed in Table 5. These primers contained an optimised Shine-Dalgarno sequence (Mega SD) [45] (link) and/or restriction sites as indicated. The sequence of the PCR products was confirmed by DNA sequencing. The M. smegmatis optimised gluc gene was cloned into pSMT3 and pSMT3M as a BamHI-HindIII fragment and into pMV306myc as an EcoRI-SalI fragment. For cloning the wild-type and optimised gluc genes into pMV306hsp the PCR products were digested with EcoRI-SalI. Plasmids containing the M. tuberculosis optimised ffluc gene were made in a similar way but cloning the EcoRI-XbaI PCR product into pUC18 first to use the restriction sites of this vector's MCS. The wild-type ffluc was cloned into pMV306hsp as a HindIII-SalI insert. Plasmids pMV306hsp+Lux, pSMT3+Lux and pSMT3M+Lux were created by cloning a 5.7-kb EcoRI-PstI blunted fragment (containing the whole lux operon) from pSB2025 into the respective expression vector. Deletion of a 0.4 kb NotI-EcoRI fragment (containing P_hsp60) from pMV306hsp+Lux and insertion of P_myc1tetO from pMV306myc produced pMV306myc+Lux. In a similar way, the reporter plasmids containing the G13 promoter were made replacing the P_hsp60 of the corresponding reporter vector with P_G13 by digestion with NotI-EcoRI. pMVhsp+LuxAB+G13+CDE was obtained by cloning the KpnI P_G13 PCR fragment into pMVhsp+Lux in front of luxC. Finally, luxCDABE from pMU1* was cloned into pMV306hsp as a 5.7-kb EcoRI PCR fragment.
Reporter strains were obtained by electroporation of reporter plasmids into M. smegmatis mc²155 or M. tuberculosis H37Rv as previously described [73] (link). Each strain was named according to the plasmid it contained. The strains transformed with an integrating vector were checked by PCR with primers amplifying the corresponding promoter and reporter gene; whereas recombinant strains with replicating vectors were confirmed by recovering the plasmid after transformation into E. coli.

Publication 2010

Cloning Vectors Deletion Mutation Deoxyribonuclease EcoRI Digestion Electroporation Escherichia coli Genes Genes, Reporter Mycobacterium tuberculosis Mycobacterium tuberculosis H37Rv Oligonucleotide Primers Operon Plasmids Strains

Mycobacterial Growth and Genetic Manipulation

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

Most recents protocols related to «Mycobacterium tuberculosis H37Rv»

Conserved MTB Epitope Targeting Antibodies

Not available on PMC !

Example 4

Composite MTB peptide vaccines induce antibodies that recognize the conserved MTB Alpha Crystallin HSP epitope (designated as TB Pep01) derived from Mycobacterium tuberculosis H37Rv (NC_000962.2).

Serum antibodies from mice 1433-1436 (see FIG. 26) immunized with 50 ug TB Pep01 CRM-conjugated vaccine demonstrated good responses to the conserved MTB epitope (alpha crystallin HSP), TB Pep01. Mouse 1435 (see FIG. 27), selected for fusion, produced hybridomas LD7 I BB2 and CA6 II GA8 that demonstrated good binding activity to TB Pep01 and TB Pep02 epitopes (FIG. 28). Monoclonal antibodies LD7 I BB2 I B9 and CA6 II GA8 I A5 (hereafter referred to as mAb LD7 and CA6) that were developed from the hybridomas not only bound to TB Pep01, but also to live M. smegmatis (see FIG. 29). Importantly, mAbs LD7 and CA6 promoted opsonophagocytic killing of mycobacteria (see FIGS. 30 and 31).

US11866463B2. Immunogenic compositions to treat and prevent microbial infections (2024-01-09). Longhorn Vaccines and Diagnostics, LLC [US]. Inventors: Luke T. Daum [US], Gerald W. Fischer [US], Clara J. Sei [US].

Patent 2024

alpha-Crystallins Antibodies Epitopes Figs Hybridomas Monoclonal Antibodies Mus Mycobacterium Mycobacterium tuberculosis H37Rv Serum Vaccines, Conjugate Vaccines, Peptide

Nanopore Sequencing and Variant Analysis of Mycobacterium tuberculosis

Nanopore raw data (fast5) were analyzed using Guppy Version 4.5.2 software (ONT) [4 (link)]. Basecalling of data was repeated using the parameter "--config dna_r9.4.1_450 bps_hac.cfg--num_callers 4 --cpu_threads_per_caller 4" then barcode recognition was conducted using the parameter "--barcode_kits EXPNBD104 EXP-NBD114" followed by trimming of sequences using the parameter" “--config configuration.cfg--trim_barcodes”. Thereafter, sequence data were counted using NanoPlot v1.28.1 [7 (link)] and variant calls were found using medaka v1.3.2 [8 ]. Finally, raw reads were mapped to the Mycobacterium tuberculosis H37Rv genomic reference sequence then trimmed reads were assembled onto the reference genome using Genomics software.

Liu Z., Yang Y., Wang Q., Wang L., Nie W, & Chu N. (2023). Diagnostic value of a nanopore sequencing assay of bronchoalveolar lavage fluid in pulmonary tuberculosis. BMC Pulmonary Medicine, 23, 77.

Publication 2023

Genome Lebistes Mycobacterium tuberculosis H37Rv Oryziinae

Evaluation of MassARRAY in Diagnosing Drug-Resistant Tuberculosis

Samples from patients with confirmed tuberculosis, including 52 bronchoalveolar lavage fluid (BALF), 42 sputum, and 128 MTB clinical isolates, were collected from Xi’an Chest Hospital. This study was approved by the Ethics Committee of Xi’an Chest Hospital [No: R2022-003-01]. Reference strains Mycobacterium tuberculosis H37Rv were obtained from the Chinese Center for Disease Control and Prevention, and CMCC95102/CMCC95103 were purchased from the National Center for Medical Culture Collections. Wild-type and mutant plasmid mixtures were synthesized by Sangon Biotech (Shanghai) Co., Ltd.
As illustrated in Figure 1, the application value of MassARRAY in the diagnosis of DR-TB was evaluated by preliminary detection performance analysis and the value of clinical application.
LOD of MassARRAY was conducted for preliminary detection performance analysis: the reference strains were diluted into 10², 10³, 10⁴, and 10⁵ CFU/mL. Reference strain DNA was extracted using three commonly used clinical nucleic acid extraction methods (Magnetic bead method, Column extraction method, and Glass bead method), and was used to detect the LOD of MassARRAY for TB identification and drug resistance gene detection. The reference strains M. tuberculosis H37Rv, CMCC95102, and CMCC95103 were used to repeat the test. Two identification sites and 25 drug-resistant genes sites were detected by MassARRAY (Table 1). The lowest concentration that can be detected is LOD.
The value of clinical application: MTB in BALF and sputum samples were detected using MassARRAY, quantitative real-time polymerase chain reaction (qPCR) and MGIT960 liquid culture (culture). Using culture as the standard, the efficacy of MassARRAY and qPCR for the detection of TB was analyzed. Mutation of drug resistance genes in MTB clinical isolates was tested using MassARRAY, HRM, and Sanger sequencing. Using sequencing as the standard, the efficacy of MassARRAY, and HRM for the detection of each drug resistance site of MTB was analyzed. Simultaneously, the mutation of drug resistance genes by the MassARRAY method was compared with the results of DST, and the genotype–phenotype relationship was analyzed. The ability of MassARRAY to discriminate mixed infections was detected using mixtures of standard strains (M. tuberculosis H37Rv) and drug-resistant clinical isolates and mixtures of wild-type and mutant plasmids. The concentration of drug-resistant gene wild plasmid and mutant plasmid was 10⁷ copies/μL, mixed in different ratios, and the ratios of mutant plasmid was 5, 10, 15, 20, and 25%. M. tuberculosis H37Rv and clinical isolates of drug-resistant strains were diluted to 10³, 10⁴, 10⁵, and 10⁶ CFU/mL, mixed in different ratios, and the ratios of clinical isolates of drug-resistant strains was 20, 40, 60, and 80%.

Yang H., Li A., Dang L., Kang T., Ren F., Ma J., Zhou Y., Yang Y., Lei J, & Zhang T. (2023). A rapid, accurate, and low-cost method for detecting Mycobacterium tuberculosis and its drug-resistant genes in pulmonary tuberculosis: Applications of MassARRAY DNA mass spectrometry. Frontiers in Microbiology, 14, 1093745.

Publication 2023

Bronchoalveolar Lavage Fluid Chest Chinese Coinfection Diagnosis Ethics Committees Genes Genotype Mutation Mycobacterium tuberculosis H37Rv Nucleic Acids Patients Pharmaceutical Preparations Phenotype Plasmids Real-Time Polymerase Chain Reaction Resistance, Drug Sputum Strains Substance Abuse Detection Tuberculosis

Real-time PCR Assay for Tuberculosis Drug Resistance

High-resolution melting curve real-time PCR experiments were performed with the SLAN-96S Real-time fluorescence quantitative PCR detection system (Zeesan Biotech, Xiamen, China), using an MTB drug resistance mutation detection kit (Fluorescent PCR melting curve method, Zeesan Biotech, Xiamen, China). Nine PCR reaction systems were performed simultaneously, in a final volume of 25 μL containing 2 μL template DNA. The reaction program was: 50°C, 60s; 95°C, 600 s; 95°C, 15 s, 70°C, 20s (reduced 1°C for each cycle), 76°C, 25 s for 13 cycles; 95°C, 15 s, 57°C, 20s, 76°C, 25 s for 42 cycles; 95°C, 120 s; 40°C, 120 s; 45°C–85°C, fluorescence signals were collected every 1°C for this period. The anti-tuberculosis drugs and their corresponding gene resistance loci are shown in Table 1. The software analyzes the differences in the shape of the melting curve between a sample and the wild-type control strain (M. tuberculosis H37Rv) by generating a difference plot curve. This plot helps with clustering samples into groups having similar melting curves; hence, sequence polymorphisms can be detected.

Publication 2023

Antitubercular Agents Fluorescence Genetic Loci Genetic Polymorphism Mutation Mycobacterium tuberculosis H37Rv Real-Time Polymerase Chain Reaction Strains Substance Abuse Detection

Sanger Sequencing of M. tuberculosis

Primers of MassArray listed in Table 1 were used for Sanger sequencing. The test was performed by Sangon Biotech (Shanghai, China), sequence analysis was performed using BLAST,¹ using M. tuberculosis H37Rv genome (NC_000962.3) as the reference sequence.

Publication 2023

Genome Mycobacterium tuberculosis H37Rv Oligonucleotide Primers Sequence Analysis

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What is Mycobacterium tuberculosis H37Rv?

What are the key differences between Mycobacterium tuberculosis H37Rv and other strains?

Mycobacterium tuberculosis H37Rv is a well-characterized and widely-used laboratory strain that exhibits consistent virulence and growth characteristics. Compared to other strains, H37Rv is known to be more virulent and better-suited for controlled research studies on tuberculosis. Its predictable behavior and genetic stability make it a preferred choice for various experimental applications in the field.

How can PubCompare.ai help optimize Mycobacterium tuberculosis H37Rv research?

PubCompare.ai allows researchers to screen protocol literature more effciently and leverage AI to pinpoint critical insights related to Mycobacterium tuberculosis H37Rv. The platform's AI-driven analysis can highlight key differences in protocol effectiveness, enabling you to choose the best option for reproducibility and accuracy in your M. tuberculosis H37Rv studies. This helps streamline your research and enhance the overall quality of your findings.

More about "Mycobacterium tuberculosis H37Rv"

Mycobacterium tuberculosis H37Rv, a widely used laboratory strain of the bacterium that causes tuberculosis, is a virulent model commonly employed in research to study the pathogenesis, immunology, and drug development for this deadly disease.
This strain, also known as M. tuberculosis H37Rv, is often cultured in Middlebrook 7H9 broth or Middlebrook 7H9 medium supplemented with Tween 80, Glycerol, and OADC enrichment, which provides the necessary nutrients for growth.
Researchers leverage the power of this model to gain insights into the complex mechanisms of tuberculosis (TB) infection and to develop more effective treatments.
By studying the behavior and characteristics of M. tuberculosis H37Rv, scientists can better understand the virulence factors, immune responses, and antibiotic susceptibility of the causative agent of TB.
The use of M. tuberculosis H37Rv in research is further enhanced by the availability of various tools and techniques, such as Middlebrook 7H10 agar for cultivation and Kanamycin for selective pressure.
Additionally, the use of Tyloxapol, a surfactant, can help improve the dispersal and aeration of the bacterial culture, contributing to more consistent and reproducible results.
PubCompare.ai, an AI-powered platform, can optimize your M. tuberculosis H37Rv research by helping you identify the most effective protocols from published literature, preprints, and patents.
By providing side-by-side comparisons of different methodologies, the tool can streamline your studies and enhance the reproducibility of your findings, ultimately advancing the understanding and treatment of this devastating disease.