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Dobos Lab: Recombinant Plasmids

Reference Standards

The production of recombinant proteins for routine use in laboratory studies is now enabled via ordering the recombinant clone for the antigen of interest. Detailed protocols and references are provided with orders so that laboratories can make proteins as needed without the delays of shipping and TBVTRM prioritization. The TBVTRM Contract will continue to generate the following recombinant proteins: ESAT-6, CFP10, Ag85a and Ag85b. These can be ordered without justification and are made available for use as reference standards. These standards may change as the needs of the research community change.


Large Scale Studies

The production of recombinant proteins for routine use in laboratory studies is now enabled via ordering the recombinant clone for the antigen of interest. Detailed protocols and references are provided with orders so that laboratories can make proteins as needed without the delays of shipping and TBVTRM prioritization. The use of recombinant proteins for in vitro studies as part of a large clinical study or trial and for structural analysis demands a purification scale that is often incompatible with the resources of most academic institutions. Thus, recombinant proteins will be made available to laboratories involved in large scale clinical trials and structural studies. Principal investigators who wish to be considered must first register with the contract and have a fully executed MTA on file. These needs will then be addressed via submission of a detailed research plan and justification. All submissions will be considered, though there may be a significant delay based on the scope of project and prior proposals in queue. Additionally, all activities in this scope of work will be considered collaborations; thus there must be a guarantee of disclosure by all parties. Please contact Dr. Karen Dobos for additional information or to submit a research proposal.


 

Reagent: Recombinant expression vectors containing Mycobacterium tuberculosis genes and protocols for generation of recombinant proteins.
Default Quantity: 1 µg plasmid DNA
Production System: All genes are amplified by PCR and cloned into pET23b or pET15b (Novagen) for expression in E. coli or pVV16 (a derivative of pMV261 allowing His-tag fusion) for expression in M. smegmatis. All proteins are His-tagged.
Notes: Recombinant proteins will only be provided as reference standards or for large scale studies (see below for more details). All other laboratories will receive plasmid DNA and detailed protocols for gene expression and protein purification.
Plasmids listed are expressed in E. coli unless otherwise stated. In general, plasmids expressed in E. coli are cloned without a signal sequence and those expressed in M. smegmatis are cloned with signal sequence (unless otherwise noted).

Table of recombinant plasmids available through the TB Vaccine Testing and Research Materials Contract
(Plasmids listed are expressed in E. coli unless otherwise stated)
Gene/Protein
Plasmid
Observed Protein Size
Expected Protein Yield*
Rv3763/19kDa/LpqH pMRLB51 19 kDa 0.5 mg
Rv2032/Acg pMRLB48 37 kDa  
Rv0733/Adk pMRLB25 29 kDa  
Rv3804/Ag85A pMRLB41 32 kDa 1 mg
Rv1886c/Ag85B pMRLB47 32 kDa 1 mg
Rv0129c/Ag85C pMRLB16 32 kDa 6 mg
Rv3841/BfrB pMRLB5 20 kDa

1 mg (high endotoxin)
0.1 mg (after ET removal)

Rv3841/BfrB pMRLB63
M. smegmatis
20 kDa  
Rv3874/CFP10 pMRLB46 11 kDa 1 mg
Rv0350/DnaK pMRLB6 70 kDa 1 mg
Rv3875/Esat6 pMRLB7 10 kDa 5 mg
Rv3019c/EsxR pMRLB67 10 kDa  
Rv1827/GarA pMRLB71 23 kDa  
Rv1837c/GlcB pMRLB8 80 kDa 0.2 mg
Rv1837c/GlcB pMRLB45
M. smegmatis
80 kDa  
Rv2220/GlnA pMRLB66 60 kDa  
Rv0440/GroEL2 pMRLB1 65 kDa 0.5 mg
Rv3418c/GroES pMRLB9 10 kDa  
Rv2031c/HspX pMRLB15 16 kDa 5 mg
Rv1484/InhA pMRLB10 29 kDa  
Rv1908c/KatG pMRLB11 81 kDa 3 mg (not clean)
0.05 mg (after cleanup)
Rv3006/LppZ pMRLB54 39 kDa  
Rv3006/Lppz pMRLB55
M. smegmatis
39 kDa  
Rv1270c/LprA pMRLB50 25 kDa  
Rv1411c/LprG pMRLB43 27 kDa 1.5 mg
Rv1411c/LprG pMRLB44
M. smegmatis
27 kDa  
Rv1860/Mpt32 pMRLB17 45 kDa 0.1 mg
Rv3803c/Mpt51 pMRLB38 31 kDa 4 mg
Rv1926c/Mpt63 pMRLB69 20 kDa  
Rv1980c/Mpt64 pMRLB12 25 kDa 3 mg
Rv0899/OmpA pMRLB42 33 kDa  
Rv0934/PhoS1/PstS pMRLB2 38 kDa 0.75 mg
Rv0652/RplL pMRLB13 13 kDa 0.25 mg
Rv0379/SecE2 pMRLB56 8 kDa  
Rv0379/SecE2 pMRLB57
M. smegmatis
8 kDa  
Rv3846/SodA pMRLB68 25 kDa  
Rv0432/SodC pMRLB60 24 kDa  
Rv0432/SodC pMRLB61
M. smegmatis
24 kDa  
Rv0432/SodC
(w/o signal sequence)
pMRLB62
M. smegmatis
24 kDa  
Rv1932/Tpx pMRLB14 17 kDa 3 mg
Rv3914/TrxC/Mpt46 pMRLB58 12 kDa  
Rv3914/TrxC/Mpt46 pMRLB59
M. smegmatis
12 kDa  
Rv0164 pMRLB70 22 kDa  
Rv0577 pMRLB24 27 kDa  
Rv1738 pMRLB49 10 kDa  
Rv1813c pMRLB53 15 kDa  
Rv2626c pMRLB28 15 kDa 8 mg
Naked Plasmid pVV16    

* Approximate yields are given if known, based on recovery of purified protein from a 1L culture.
Note: This list is updated on a continual basis as new recombinant products are generated.

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