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pNam vector (V004362)

Basic Vector Information

Vector Name:
pNam
Antibiotic Resistance:
Ampicillin
Length:
6278 bp
Type:
T7 Expression vector
Replication origin:
pSC101 ori
Source/Author:
Hoang TT, Stern RJ, McNeil MR, Schweizer HP.

pNam vector Vector Map

pNam6278 bp30060090012001500180021002400270030003300360039004200450048005100540057006000AmpRAmpR promoterT7 terminatorCBDSce VMA intein 3' regionSce VMA intein 5' regionmultiple cloning siteRBSlac operatorT7 promoterlacI promoterlacICAP binding sitepSC101 oriRep101

Plasmid Resuspension Protocol:

1. Centrifuge at 5,000×g for 5 min.

2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.

3. Close the tube and incubate for 10 minutes at room temperature.

4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.

5.Store the plasmid at -20 ℃.

pNam vector Sequence

Copy Sequence

Download GeneBank File(.gb)

LOCUS       40924_32988        6278 bp DNA     circular SYN 18-DEC-2018
DEFINITION  T7 Expression vector pNam, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6278)
  AUTHORS   Hoang TT, Stern RJ, McNeil MR, Schweizer HP.
  TITLE     Construction and use of low-copy number T7 expression vectors for 
            purification of problem proteins: affinity purification of 
            Mycobacterium tuberculosis RmlD and Pseudomonas aeruginosa LasI and 
            RhlI proteins, and functional analysis of RhlI
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6278)
  AUTHORS   Hoang TT, Schweizer HP.
  TITLE     Direct Submission
  JOURNAL   Submitted (30-APR-1999) Microbiology, Colorado State University, 
            Fort Collins, CO 80523, USA
REFERENCE   3  (bases 1 to 6278)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 6278)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (30-APR-1999) Microbiology, Colorado State University, Fort Collins,
            CO 80523, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6278
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(147..1004)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERSPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(1005..1109)
                     /label=AmpR promoter
     terminator      complement(1169..1216)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(1295..1450)
                     /codon_start=1
                     /label=CBD
                     /note="chitin binding domain from chitinase A1 (Watanabe et
                     al., 1994)"
                     /translation="TTNPGVSAWQVNTAYTAGQLVTYNGKTYKCLQPHTSLAGWEPSNV
                     PALWQLQ"
     CDS             complement(1490..2023)
                     /codon_start=1
                     /label=Sce VMA intein 3' region
                     /note="modified 3' region of the intein from the yeast Vma1
                     subunit of the vacuolar ATPase (Chong et al., 1998)"
                     /translation="GIRNNLNTENPLWDAIVGLGFLKDGVKNIPSFLSTDNIGTRETFL
                     AGLIDSDGYVTDEHGIKATIKTIHTSVRDGLVSLARSLGLVVSVNAEPAKVDMNVTKHK
                     ISYAIYMSGGDVLLNVLSKCAGSKKFRPAPAAAFARECRGFYFELQELKEDDYYGITLS
                     DDSDHQFLLGSQVVV"
     CDS             complement(2027..2845)
                     /codon_start=1
                     /label=Sce VMA intein 5' region
                     /note="5' region of the intein from the yeast Vma1 subunit
                     of the vacuolar ATPase (Chong et al., 1998)"
                     /translation="CFAKGTNVLMADGSIECIENIEVGNKVMGKDGRPREVIKLPRGRE
                     TMYSVVQKSQHRAHKSDSSREVPELLKFTCNATHELVVRTPRSVRRLSRTIKGVEYFEV
                     ITFEMGQKKAPDGRIVELVKEVSKSYPISEGPERANELVESYRKASNKAYFEWTIEARD
                     LSLLGSHVRKATYQTYAPILYENDHFFDYMQKSKFHLTIEGPKVLAYLLGLWIGDGLSD
                     RATFSVDSRDTSLMERVTEYAEKLNLCAEYKDRKEPQVAKTVNLYSKVVRG"
     misc_feature    2846..2899
                     /label=multiple cloning site
                     /note="multiple cloning site"
     RBS             complement(2904..2926)
                     /label=RBS
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    complement(2941..2965)
                     /label=lac operator
                     /note="The lac repressor binds to the lac operator to
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(2966..2984)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        3297..3374
                     /label=lacI promoter
     CDS             3375..4454
                     /codon_start=1
                     /label=lacI
                     /note="lac repressor"
                     /translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     protein_bind    4470..4491
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      4956..5178
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     CDS             5226..6173
                     /codon_start=1
                     /label=Rep101
                     /note="RepA protein needed for replication with the pSC101 
                     origin"
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"

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