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  JOURNAL   Unpublished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            "Unpublished"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"
ORIGIN
        1 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
       61 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
      121 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
      181 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
      241 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
      301 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
      361 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
      421 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
      481 tcgtcgtttg gtatggcttc attcagctcc ggttcccagc gatcaaggcg agttacatga
      541 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
      601 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
      661 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
      721 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa cacgggataa taccgcgcca
      781 catagcagaa ctttaaaagt gctcatcatt ggagaacgtt cttcggggcg aaaactctca
      841 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
      901 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
      961 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     1021 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     1081 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgta
     1141 gttatccgga tatagttcct cctttcagca aaaaacccct caagacccgt ttagaggccc
     1201 caaggggtta tgctagttat tgctcagcgg tggcagcagc caactcagct tcctttcggg
     1261 ctttgttagc agccggatcc ccttcctgca gtcattgaag ctgccacaag gcaggaacgt
     1321 tggatggttc ccatcctgcc aaggaggtgt ggggctgcaa acatttatac gtcttgccgt
     1381 tatatgtgac caattgtccc gcagtataag ctgtgttgac ctgccaagcg gatacaccag
     1441 gatttgtcgt gaggcctgag ttcagaccgg tcaggccacc gcatgcatgg acgacaacct
     1501 gggatccaag caaaaactga tgatcagaat catcagataa agtaatccca taataatcgt
     1561 cttccttcaa ttcttgtaac tcgaaataaa atccgcggca ctcacgtgca aaagcagcgg
     1621 cgggagcagg cctgaatttt ttagagccgg cacacttcga aagaacgtta agcaaaacat
     1681 ctccaccaga catataaata gcataactaa ttttatgttt ggtgacattc atgtcaacct
     1741 tagcaggttc tgcgttaacc gagactacta agcctaaaga acgagcaagg gaaaccaaac
     1801 catctctgac agaagtatga attgtcttta ttgttgcttt aataccatgc tcatcagtaa
     1861 catagccatc agaatcaatt agaccagcaa gaaatgtttc acgagtaccg atattgtccg
     1921 tagacaagaa agaaggaata tttttgacac cgtccttcaa gaatcctaag ccaacaatag
     1981 cgtcccataa tggattctca gtattaagat tattgcgaat accattacct ctgacaactt
     2041 tagagtacaa attaacagtt ttggcaactt gtggttcttt tctgtcctta tactcggcgc
     2101 acaaattcaa cttttcagca tattcagtaa cacgttccat caaagaagta tctctggaat
     2161 caaccgaaaa agttgccctg tcagacaatc catcaccaat ccataaacca agtaaataag
     2221 caagtacttt tggaccttca atggtgagat gaaacttact tttttgcatg tagtcgaaaa
     2281 agtggtcatt ctcataaaga attggagcgt aagtctggta ggtagcttta cgaacatggg
     2341 aacccaacag agaaagatct ctggcctcaa tagtccactc aaaataagct ttatttgaag
     2401 cctttctata ggattctact aattcgttgg ctctctcagg cccctcagat attgggtagc
     2461 tctttgaaac ttccttgaca agctcaacaa ttctaccgtc gggggctttc ttttggccca
     2521 tctcaaaagt aataacttca aaatattcga cacccttaat ggtacgagac aaacggcgga
     2581 cactacgagg tgttctaaca accaactcat gggtcgcatt acacgtaaac ttgagtaatt
     2641 ctggcacttc acgacttgag tcacttttgt gggctctgtg ctgacttttc tgcacgacgc
     2701 tgtacatagt ttctcttcct ctgggcaatt taattacctc acgaggtctg ccatctttac
     2761 ccatgacctt attaccaacc tcaatgtttt caatacattc aatagaccca tccgccatta
     2821 aaacattggt acccttggca aagcaggaag agccctcgag gaattcgcgg ccgccgtcga
     2881 ctcgcgagct agccatatgt atatctcctt cttaaagtta aacaaaatta tttctagagg
     2941 ggaattgtta tccgctcaca attcccctat agtgagtcgt attaatttcg cgggatcgag
     3001 atcgatctcg atcctctacg ccggacgcat cgtggccggc atcaccggcg ccacaggtgc
     3061 ggttgctggc gcctatatcg ccgacatcac cgatggggaa gatcgggctc gccacttcgg
     3121 gctcatgagc gcttgtttcg gcgtgggtat ggtggcaggc cccgtggccg ggggactgtt
     3181 gggcgccatc tccttgcatg caccattcct tgcggcggcg gtgctcaacg gcctcaacct
     3241 actactgggc tgcttcctaa tgcaggagtc gcataaggga gagcgtcgag atcccggaca
     3301 ccatcgaatg gcgcaaaacc tttcgcggta tggcatgata gcgcccggaa gagagtcaat
     3361 tcagggtggt gaatgtgaaa ccagtaacgt tatacgatgt cgcagagtat gccggtgtct
     3421 cttatcagac cgtttcccgc gtggtgaacc aggccagcca cgtttctgcg aaaacgcggg
     3481 aaaaagtgga agcggcgatg gcggagctga attacattcc caaccgcgtg gcacaacaac
     3541 tggcgggcaa acagtcgttg ctgattggcg ttgccacctc cagtctggcc ctgcacgcgc
     3601 cgtcgcaaat tgtcgcggcg attaaatctc gcgccgatca actgggtgcc agcgtggtgg
     3661 tgtcgatggt agaacgaagc ggcgtcgaag cctgtaaagc ggcggtgcac aatcttctcg
     3721 cgcaacgcgt cagtgggctg atcattaact atccgctgga tgaccaggat gccattgctg
     3781 tggaagctgc ctgcactaat gttccggcgt tatttcttga tgtctctgac cagacaccca
     3841 tcaacagtat tattttctcc catgaagacg gtacgcgact gggcgtggag catctggtcg
     3901 cattgggtca ccagcaaatc gcgctgttag cgggcccatt aagttctgtc tcggcgcgtc
     3961 tgcgtctggc tggctggcat aaatatctca ctcgcaatca aattcagccg atagcggaac
     4021 gggaaggcga ctggagtgcc atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg
     4081 gcatcgttcc cactgcgatg ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg
     4141 ccattaccga gtccgggctg cgcgttggtg cggatatctc ggtagtggga tacgacgata
     4201 ccgaagacag ctcatgttat atcccgccgt taaccaccat caaacaggat tttcgcctgc
     4261 tggggcaaac cagcgtggac cgcttgctgc aactctctca gggccaggcg gtgaagggca
     4321 atcagctgtt gcccgtctca ctggtgaaaa gaaaaaccac cctggcgccc aatacgcaaa
     4381 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
     4441 tggaaagcgg gcagtgagcg caacgcaatt aatgtaagtt agctcactca ttaggcaccg
     4501 ggatctcgac cgatgccctt gagagccttc aacccagtca gctccttccg gtgggcgcgg
     4561 ggcatgacta tcgtcgccgc acttatgact gtcttcttta tcatgcaact cgtaggacag
     4621 gtgccggcag cgctctgggt cattttcggc gaggaccgct ttcgctggag cgcgacgatg
     4681 atcggcctgt cgcttgcggt attcggaatc ttgcacgccc tcgctcaagc cttcgtcact
     4741 ggtcccgcca ccaaacgttt cggcgagaag caggccatta tcgccggcat ggcggccgac
     4801 gcgctgggct acgtcttgct ggcgttcgat tcagcgattt gcccgagctt gcgagggtgc
     4861 tacttaagcc tttagggttt taaggtctgt tttgtagagg agcaaacagc gtttgcgaca
     4921 tccttttgta atactgcgga actgactaaa gtagtgagtt atacacaggg ctgggatcta
     4981 ttctttttat ctttttttat tctttcttta ttctataaat tataaccact tgaatataaa
     5041 caaaaaaaac acacaaaggt ctagcggaat ttacagaggg tctagcagaa tttacaagtt
     5101 ttccagcaaa ggtctagcag aatttacaga tacccacaac tcaaaggaaa aggactagta
     5161 attatcattg actagcccat ctcaattggt atagtgatta aaatcaccta gaccaattga
     5221 gatgtatgtc tgaattagtt gttttcaaag caaatgaact agcgattagt cgctatgact
     5281 taacggagca tgaaaccaag ctaattttat gctgtgtggc actactcaac cccacgattg
     5341 aaaaccctac aaggaaagaa cggacggtat cgttcactta taaccaatac gctcagatga
     5401 tgaacatcag tagggaaaat gcttatggtg tattagctaa agcaaccaga gagctgatga
     5461 cgagaactgt ggaaatcagg aatcctttgg ttaaaggctt tgagattttc cagtggacaa
     5521 actatgccaa gttctcaagc gaaaaattag aattagtttt tagtgaagag atattgcctt
     5581 atcttttcca gttaaaaaaa ttcataaaat ataatctgga acatgttaag tcttttgaaa
     5641 acaaatactc tatgaggatt tatgagtggt tattaaaaga actaacacaa aagaaaactc
     5701 acaaggcaaa tatagagatt agccttgatg aatttaagtt catgttaatg cttgaaaata
     5761 actaccatga gtttaaaagg cttaaccaat gggttttgaa accaataagt aaagatttaa
     5821 acacttacag caatatgaaa ttggtggttg ataagcgagg ccgcccgact gatacgttga
     5881 ttttccaagt tgaactagat agacaaatgg atctcgtaac cgaacttgag aacaaccaga
     5941 taaaaatgaa tggtgacaaa ataccaacaa ccattacatc agattcctac ctacataacg
     6001 gactaagaaa aacactacac gatgctttaa ctgcaaaaat tcagctcacc agttttgagg
     6061 caaaattttt gagtgacatg caaagtaagt atgatctcaa tggttcgttc tcatggctca
     6121 cgcaaaaaca acgaaccaca ctagagaaca tactggctaa atacggaagg atctgaggtt
     6181 cttatggctc ttgtatctat cagtgaagca tcaagactaa caaacaaaag tagaacaact
     6241 gttcaccgtt acatatcaaa gggaaaactg tccataga
//
