LOCUS       40924_39783        6835 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pSFFLAG-CTC, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6835)
  AUTHORS   Couch RD, Seidle HF, Parry RJ.
  TITLE     Construction of Expression Vectors to Produce Affinity Tagged 
            Proteins in Pseudomonas
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6835)
  AUTHORS   Couch RD, Seidle HF, Parry RJ.
  TITLE     Direct Submission
  JOURNAL   Submitted (19-DEC-2001) Chemistry, Rice University, 1900 Rice Blvd 
            MS60, Houston, TX 77005, USA
REFERENCE   3  (bases 1 to 6835)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 6835)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (19-DEC-2001) Chemistry, Rice University, 1900 Rice Blvd MS60, 
            Houston, TX 77005, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6835
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(69..242)
                     /codon_start=1
                     /label=lacZ-alpha
                     /note="LacZ-alpha fragment of beta-galactosidase"
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGE"
     protein_bind    complement(262..278)
                     /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(286..316)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(331..352)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             complement(368..1447)
                     /codon_start=1
                     /label=lacI
                     /note="lac repressor"
                     /translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        complement(1448..1525)
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the
                     promoter boosts expression of the lacI gene about 10-fold."
     promoter        2066..2094
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and
                     lac UV5 promoters"
     protein_bind    2102..2118
                     /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)."
     CDS             2177..2200
                     /codon_start=1
                     /label=FLAG
                     /note="FLAG(R) epitope tag, followed by an enterokinase
                     cleavage site"
                     /translation="DYKDDDDK"
     terminator      2411..2497
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     terminator      2589..2616
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     promoter        2635..2726
                     /label=AmpR promoter
     CDS             2727..3584
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      3758..4346
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     rep_origin      complement(4958..5309)
                     /direction=LEFT
                     /label=pRO1600 oriV
                     /note="broad-host-range origin of replication from
                     Pseudomonas aeruginosa plasmid pRO1600; requires the 
                     pRO1600 Rep protein for replication (West et al., 1994)"
     CDS             5323..6153
                     /codon_start=1
                     /label=pRO1600 Rep
                     /note="replication protein for the broad-host-range plasmid
                     pRO1600 from Pseudomonas aeruginosa"
                     /translation="VASPPMVYKSNALVEAAYRLSVQEQRIVLACISQVKRSEPVTDEV
                     MYSVTAEDIATMAGVPIESSYNQLKEAALRLKRREVRLTQEPNGKGKRPSVMITGWVQT
                     IIYREGEGRVELRFTKDMLPYLTELTKQFTKYALADVAKMDSTHAIRLYELLMQWDSIG
                     QREIEIDQLRKWFQLEGRYPSIKDFKLRVLDPAVTQINEHSPLQVEWAQRKTGRKVTHL
                     LFSFGPKKPAKAVGKAPAKRKAGKISDAEIAKQARPGETWEAARARLTQMPLDLA"
     rep_origin      6401..6829
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
ORIGIN
        1 tcaggaagat cgcactccag ccagctttcc ggcaccgctt ctggtgccgg aaaccaggca
       61 aagcgccatt cgccattcag gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct
      121 tcgctattac gccagctggc gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg
      181 ccagggtttt cccagtcacg acgttgtaaa acgacggcca gtgaatccgt aatcatggtc
      241 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
      301 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taacttacat taattgcgtt
      361 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
      421 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgccag ggtggttttt cttttcacca
      481 gtgagacggg caacagctga ttgcccttca ccgcctggcc ctgagagagt tgcagcaagc
      541 ggtccacgct ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt aacggcggga
      601 tataacatga gctgtcttcg gtatcgtcgt atcccactac cgagatatcc gcaccaacgc
      661 gcagcccgga ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg ttggcaacca
      721 gcatcgcagt gggaacgatg ccctcattca gcatttgcat ggtttgttga aaaccggaca
      781 tggcactcca gtcgccttcc cgttccgcta tcggctgaat ttgattgcga gtgagatatt
      841 tatgccagcc agccagacgc agacgcgccg agacagaact taatgggccc gctaacagcg
      901 cgatttgctg gtgacccaat gcgaccagat gctccacgcc cagtcgcgta ccgtcttcat
      961 gggagaaaat aatactgttg atgggtgtct ggtcagagac atcaagaaat aacgccggaa
     1021 cattagtgca ggcagcttcc acagcaatgg catcctggtc atccagcgga tagttaatga
     1081 tcagcccact gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc
     1141 cgcttcgttc taccatcgac accaccacgc tggcacccag ttgatcggcg cgagatttaa
     1201 tcgccgcgac aatttgcgac ggcgcgtgca gggccagact ggaggtggca acgccaatca
     1261 gcaacgactg tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg
     1321 ccatcgccgc ttccactttt tcccgcgttt tcgcagaaac gtggctggcc tggttcacca
     1381 cgcgggaaac ggtctgataa gagacaccgg catactctgc gacatcgtat aacgttactg
     1441 gtttcacatt caccaccctg aattgactct cttccgggcg ctatcatgcc ataccgcgaa
     1501 aggttttgca ccattccatg gtgtcggaat tgctgcaggt cgagggggtc atggctgcgc
     1561 cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg
     1621 cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat
     1681 caccgaaacg cgcgaggcag gatggcgccc aacagtcccc cggccacggg gcctgccacc
     1741 atacccacgc cgaaacaagc gctcatgagc ccgaagtggc gagcccgatc ttccccatcg
     1801 gtgatgtcgg cgatataggc gccagcaacc gcacctgtgg cgccggtgat gccggccacg
     1861 atgcgtccgg cgtagaggat ccgggcttat cgactgcacg gtgcaccaat gcttctggcg
     1921 tcaggcagcc atcggaagct gtggtatggc tgtgcaggtc gtaaatcact gcataattcg
     1981 tgtcgctcaa ggcgcactcc cgttctggat aatgtttttt gcgccgacat cataacggtt
     2041 ctggcaaata ttctgaaatg agctgttgac aattaatcat cggctcgtat aatgtgtgga
     2101 attgtgagcg gataacaatt tcacacagga gatatcatat gaagcttctc gagaattccc
     2161 gggtaccaga tctgtcgact acaaggacga tgacgacaag tgaagatcga tctctcgatc
     2221 gagtgagaga agattttcag cctgatacag attaaatcag aacgcagaag cggtctgata
     2281 aaacagaatt tgcctggcgg cagtagcgcg gtggtcccac ctgaccccat gccgaactca
     2341 gaagtgaaac gccgtagcgc cgatggtagt gtggggtctc cccatgcgag agtagggaac
     2401 tgccaggcat caaataaaac gaaaggctca gtcgaaagac tgggcctttc gttttatctg
     2461 ttgtttgtcg gtgaacgctc tcctgagtag gacaaatccg ccgggagcgg atttgaacgt
     2521 tgcgaagcaa cggcccggag ggtggcgggc aggacgcccg ccataaactg ccaggcatca
     2581 aattaagcag aaggccatcc tgacggatgg cctttttgcg tttctacaaa ctcttttgtt
     2641 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     2701 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     2761 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     2821 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     2881 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     2941 ttctgctatg tggcgcggta ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc
     3001 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     3061 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     3121 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     3181 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     3241 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat
     3301 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     3361 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     3421 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     3481 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     3541 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     3601 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     3661 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     3721 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     3781 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     3841 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     3901 ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     3961 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     4021 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     4081 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     4141 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     4201 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     4261 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     4321 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     4381 ccttttgctg gccttttgct cacatgtcca cttctgcgct cggcccttcc ggctggctgg
     4441 tttattgctg ataaatctgg agccggtgag cgtggatctc gcggtatcat tgcagcactg
     4501 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     4561 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     4621 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     4681 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     4741 ttttcgttcc actgagcgtc agaccccaat tacacgccac tggctgtgct tgctggggtg
     4801 acggtggcaa cggtggcggc cttgctgggc tatcgcgttg gaaagaaacg agggaaaggg
     4861 gactgataaa ccggtcttag cccctcccct tggtgtccaa ccgctctgta ggcctctcag
     4921 gcgccgctgg tgccgctggt tggacgccaa gggtgaatcc gcctcgatac cctgattact
     4981 cgcttcctgc gccctctcag gcggcgatag gggactggta aaacggggat tgcccagacg
     5041 cctcccccgc cccttcaggg gcacaaatgc ggccccaacg gggccacgta gtggtgcgtt
     5101 ttttgcgttt ccaccctttt cttccttttc ccttttaaac cttttaggac gtctacaggc
     5161 cacgtaatcc gtggcctgta gagtttaaaa agggacggat ttgttgccat taagggacgg
     5221 atttgttgtt aagaagggac ggatttgttg ttgtaaaggg acggatttgt tgtattgtgg
     5281 gacgcagata cagtgtcccc ttatacacaa ggaatgtcga acgtggcctc acccccaatg
     5341 gtttacaaaa gcaatgccct ggtcgaggcc gcgtatcgcc tcagtgttca ggaacagcgg
     5401 atcgttctgg cctgtattag ccaggtgaag aggagcgagc ctgtcaccga tgaagtgatg
     5461 tattcagtga cggcggagga catagcgacg atggcgggtg tccctatcga atcttcctac
     5521 aaccagctca aagaagcggc cctgcgcctg aaacggcggg aagtccggtt aacccaagag
     5581 cccaatggca aggggaaaag accgagtgtg atgattaccg gctgggtgca aacaatcatc
     5641 taccgggagg gtgagggccg tgtagaactc aggttcacca aagacatgct gccgtacctg
     5701 acggaactca ccaaacagtt caccaaatac gccttggctg acgtggccaa gatggacagc
     5761 acccacgcga tcaggcttta cgagctgctc atgcaatggg acagcatcgg ccagcgcgaa
     5821 atagaaattg accagctgcg aaagtggttt caactggaag gccggtatcc ctcgatcaag
     5881 gacttcaagt tgcgagtgct tgatccagcc gtgacgcaga tcaacgagca cagcccgcta
     5941 caggtggagt gggcgcagcg aaagaccggg cgcaaggtca cacatctgtt gttcagtttt
     6001 ggaccgaaga agcccgccaa ggcggtgggt aaggccccag cgaagcgcaa ggccgggaag
     6061 atttcagatg ctgagatcgc gaaacaggct cgccctggtg agacatggga agcggcccgc
     6121 gctcgactaa cccagatgcc gctggatctg gcctagaggc cgtggccacc acggcccggc
     6181 ctgcctttca ggctgcgcaa ctgttgggaa gggcctcttt cctgcgttat cccctgattc
     6241 tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac
     6301 cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt attttctcct
     6361 tacgcatctg tgcggtattt cacaccgcat acagatcctg acgcgccctg tagcggcgca
     6421 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
     6481 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     6541 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     6601 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     6661 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     6721 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
     6781 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt taaca
//
