LOCUS       40924_39827        7834 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pSFV4, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7834)
  AUTHORS   Loppnau PA, Graslund S, Arrowsmith CH.
  TITLE     A pipeline to generate recombinant antibodies to human target 
            proteins involved in epigenetic mechanisms
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7834)
  AUTHORS   Loppnau PA, Graslund S, Arrowsmith CH.
  TITLE     Direct Submission
  JOURNAL   Submitted (14-NOV-2012) Structural Genomics Consortium, University 
            of Toronto, MaRS Centre, South tower, 101 College St, Room 700, 
            Toronto, Ontario M5G 1L7, Canada
REFERENCE   3  (bases 1 to 7834)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 7834)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (14-NOV-2012) Structural Genomics Consortium, University of Toronto,
            MaRS Centre, South tower, 101 College St, Room 700, Toronto, Ontario
            M5G 1L7, Canada"
COMMENT     SGRef: number: 3; type: "Journal Article"
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..7834
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      12..465
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        492..596
                     /label=AmpR promoter
     CDS             597..1454
                     /label=AmpR
                     /note="beta-lactamase"
     rep_origin      1628..2216
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    complement(2402..2544)
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(2953..3126)
                     /label=lacZ-alpha
                     /note="LacZ-alpha fragment of beta-galactosidase"
     protein_bind    complement(3146..3162)
                     /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(3170..3200)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(3215..3236)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             complement(3252..4331)
                     /label=lacI
                     /note="lac repressor"
     promoter        complement(4332..4409)
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the
                     promoter boosts expression of the lacI gene about 10-fold."
     protein_bind    4461..4482
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        4497..4527
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an 'up' mutation"
     protein_bind    4535..4551
                     /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        4606..4634
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and
                     lac UV5 promoters"
     protein_bind    4640..4660
                     /label=lac repressor protein binding site
                     /bound_moiety="lac repressor protein"
     protein_bind    4642..4658
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /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)."
     regulatory      4701..4729
                     /label=Ptac
                     /note="Ptac"
                     /regulatory_class="promoter"
     promoter        4701..4729
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and
                     lac UV5 promoters"
     regulatory      4701..4706
                     /label=derived from trp promoter
                     /note="derived from trp promoter"
                     /regulatory_class="minus_35_signal"
     regulatory      4723..4729
                     /label=derived from lacUV5 promoter
                     /note="derived from lacUV5 promoter"
                     /regulatory_class="minus_10_signal"
     protein_bind    4735..4755
                     /label=lac repressor protein binding site
                     /bound_moiety="lac repressor protein"
     protein_bind    4737..4753
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /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)."
     sig_peptide     4796..4861
                     /label=pelB signal sequence
                     /note="leader peptide for secretion"
     misc_feature    4862..5127
                     /note="partial sequence of FAB light chain constant region"
     misc_feature    5113..5127
                     /label=5'-In-Fusion cloning site
                     /note="5'-In-Fusion cloning site"
     protein_bind    complement(5143..5176)
                     /label=loxP
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        5191..5636
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             5637..7055
                     /label=SacB
                     /note="secreted levansucrase that renders bacterial growth 
                     sensitive to sucrose"
     misc_feature    7120..7418
                     /note="partial coding sequence of FAB heavy chain constant 
                     region"
     misc_feature    7120..7134
                     /label=3'-In-Fusion cloning site
                     /note="3'-In-Fusion cloning site"
     misc_feature    7419..7463
                     /label=C-terminal of FAB heavy chain with AviTag addition
                     /note="C-terminal of FAB heavy chain with AviTag addition"
     CDS             7485..7529
                     /label=AviTag(TM)
                     /note="peptide tag that allows for enzymatic biotinylation"
     misc_feature    7543..7587
                     /note="alternate C-terminal of FAB heavy chain without tag 
                     addition"
     misc_feature    7606..7644
                     /note="alternate C-terminal of FAB heavy chain with hexa 
                     histidine-tag"
     CDS             7654..7671
                     /label=6xHis
                     /note="6xHis affinity tag"
     regulatory      7716..7743
                     /label=trp operon transcription termination sequence
                     /note="trp operon transcription termination sequence"
                     /regulatory_class="terminator"
ORIGIN
        1 gcaaccatag tacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
       61 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
      121 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
      181 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgatttgg gtgatggttc
      241 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
      301 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cgggctattc
      361 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
      421 acaaaaattt aacgcgaatt ttaacaaaat attaacgttt acaatcaggt ggcacttttc
      481 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
      541 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
      601 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
      661 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
      721 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
      781 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgtg
      841 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
      901 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
      961 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     1021 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     1081 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     1141 cagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     1201 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     1261 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtcccgcg
     1321 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     1381 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     1441 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     1501 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     1561 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     1621 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     1681 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     1741 ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc
     1801 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     1861 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     1921 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     1981 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     2041 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     2101 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     2161 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     2221 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     2281 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     2341 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     2401 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atatatggtg
     2461 cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatacac tccgctatcg
     2521 ctacgtgact gggtcatggc tgcgccccga cacccgccaa cacccgctga cgcgccctga
     2581 cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc
     2641 atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga ggcagaacgc catcaaaaat
     2701 aattcgcgtc tggccttcct gtagccagct ttcatcaaca ttaaatgtga gcgagtaaca
     2761 acccgtcgga ttctccgtgg gaacaaacgg cggattgacc gtaatgggat aggttacgtt
     2821 ggtgtagatg ggcgcatcgt aaccgtgcat ctgccagttt gaggggacga cgacagtatc
     2881 ggcctcagga agatcgcact ccagccagct ttccggcacc gcttctggtg ccggaaacca
     2941 ggcaaagcgc cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc
     3001 ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct gcaaggcgat taagttgggt
     3061 aacgccaggg ttttcccagt cacgacgttg taaaacgacg gccagtgaat ccgtaatcat
     3121 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     3181 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     3241 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     3301 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggtggt ttttcttttc
     3361 accagtgaga cgggcaacag ctgattgccc ttcaccgcct ggccctgaga gagttgcagc
     3421 aagcggtcca cgctggtttg ccccagcagg cgaaaatcct gtttgatggt ggttgacggc
     3481 gggatataac atgagctgtc ttcggtatcg tcgtatccca ctaccgagat atccgcacca
     3541 acgcgcagcc cggactcggt aatggcgcgc attgcgccca gcgccatctg atcgttggca
     3601 accagcatcg cagtgggaac gatgccctca ttcagcattt gcatggtttg ttgaaaaccg
     3661 gacatggcac tccagtcgcc ttcccgttcc gctatcggct gaatttgatt gcgagtgaga
     3721 tatttatgcc agccagccag acgcagacgc gccgagacag aacttaatgg gcccgctaac
     3781 agcgcgattt gctggtgacc caatgcgacc agatgctcca cgcccagtcg cgtaccgtct
     3841 tcatgggaga aaataatact gttgatgggt gtctggtcag agacatcaag aaataacgcc
     3901 ggaacattag tgcaggcagc ttccacagca atggcatcct ggtcatccag cggatagtta
     3961 atgatcagcc cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg
     4021 acgccgcttc gttctaccat cgacaccacc acgctggcac ccagttgatc ggcgcgagat
     4081 ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca gactggaggt ggcaacgcca
     4141 atcagcaacg actgtttgcc cgccagttgt tgtgccacgc ggttgggaat gtaattcagc
     4201 tccgccatcg ccgcttccac tttttcccgc gttttcgcag aaacgtggct ggcctggttc
     4261 accacgcggg aaacggtctg ataagagaca ccggcatact ctgcgacatc gtataacgtt
     4321 actggtttca cattcaccac cctgaattga ctctcttccg ggcgctatca tgccataccg
     4381 cgaaaggttt tgcaccattc gatggtgtcc tggcacgaca ggtttcccga ctggaaagcg
     4441 ggcagtgagc gcaacgcaat taatgtaagt tagctcactc attaggcacc ccaggcttta
     4501 cactttatgc ttccggctcg tataatgtgt ggaattgtga gcggataaca atttcacaca
     4561 ggaaacagga tcgatccatc gatgagctta ctccccatcc ccctgttgac aattaatcat
     4621 cggctcgtat aatgtgtgga attgtgagcg gataacaatt tcacacagga aacaggatca
     4681 gcttactccc catccccctg ttgacaatta atcatcggct cgtataatgt gtggaattgt
     4741 gagcggataa caatttcaca caggaaacag gatccatcga tgcttaggag gtcatatgaa
     4801 atccctattg cctacggcag ccgctggatt gttattactc gcggcccagc cggccatggc
     4861 gtccgatatc cagatgaccc agtccccgag ctccctgtcc gcctctgtgg gcgatagggt
     4921 caccatcacc tgccgtgcca gtcagtccgt gtccagcgct gtagcctggt atcaacagaa
     4981 accaggaaaa gctccgaagc ttctgattta ctcggcatcc agcctctact ctggagtccc
     5041 ttctcgcttc tctggtagcc gttccgggac ggatttcact ctgaccatca gcagtctgca
     5101 gccggaagac ttcgcaactt attactgcat gctgaaagat ccataacttc gtatagcata
     5161 cattatacga agttatgcgg ccgcgacgtc cacatatacc tgccgttcac tattatttag
     5221 tgaaatgaga tattatgata ttttctgaat tgtgattaaa aaggcaactt tatgcccatg
     5281 caacagaaac tataaaaaat acagagaatg aaaagaaaca gatagatttt ttagttcttt
     5341 aggcccgtag tctgcaaatc cttttatgat tttctatcaa acaaaagagg aaaatagacc
     5401 agttgcaatc caaacgagag tctaatagaa tgaggtcgaa aagtaaatcg cgcgggtttg
     5461 ttactgataa agcaggcaag acctaaaatg tgtaaagggc aaagtgtata ctttggcgtc
     5521 accccttaca tattttaggt ctttttttat tgtgcgtaac taacttgcca tcttcaaaca
     5581 ggagggctgg aagaagcaga ccgctaacac agtacataaa aaaggagaca tgaacgatga
     5641 acatcaaaaa gtttgcaaaa caagcaacag tattaacctt tactaccgca ctgctggcag
     5701 gaggcgcaac tcaagcgttt gcgaaagaaa cgaaccaaaa gccatataag gaaacatacg
     5761 gcatttccca tattacacgc catgatatgc tgcaaatccc tgaacagcaa aaaaatgaaa
     5821 aatatcaagt tcctgagttc gattcgtcca caattaaaaa tatctcttct gcaaaaggcc
     5881 tggacgtttg ggacagctgg ccattacaaa acgctgacgg cactgtcgca aactatcacg
     5941 gctaccacat cgtctttgca ttagccggag atcctaaaaa tgcggatgac acatcgattt
     6001 acatgttcta tcaaaaagtc ggcgaaactt ctattgacag ctggaaaaac gctggccgcg
     6061 tctttaaaga cagcgacaaa ttcgatgcaa atgattctat cctaaaagac caaacacaag
     6121 aatggtcagg ttcagccaca tttacatctg acggaaaaat ccgtttattc tacactgatt
     6181 tctccggtaa acattacggc aaacaaacac tgacaactgc acaagttaac gtatcagcat
     6241 cagacagctc tttgaacatc aacggtgtag aggattataa atcaatcttt gacggtgacg
     6301 gaaaaacgta tcaaaatgta cagcagttca tcgatgaagg caactacagc tcaggcgaca
     6361 accatacgct gagagatcct cactacgtag aagataaagg ccacaaatac ttagtatttg
     6421 aagcaaacac tggaactgaa gatggctacc aaggcgaaga atctttattt aacaaagcat
     6481 actatggcaa aagcacatca ttcttccgtc aagaaagtca aaaacttctg caaagcgata
     6541 aaaaacgcac ggctgagtta gcaaacggcg ctctcggtat gattgagcta aacgatgatt
     6601 acacactgaa aaaagtgatg aaaccgctga ttgcatctaa cacagtaaca gatgaaattg
     6661 aacgcgcgaa cgtctttaaa atgaacggca aatggtacct gttcactgac tcccgcggat
     6721 caaaaatgac gattgacggc attacgtcta acgatattta catgcttggt tatgtttcta
     6781 attctttaac tggcccatac aagccgctga acaaaactgg ccttgtgtta aaaatggatc
     6841 ttgatcctaa cgatgtaacc tttacttact cacacttcgc tgtacctcaa gcgaaaggaa
     6901 acaatgtcgt gattacaagc tatatgacaa acagaggatt ctacgcagac aaacaatcaa
     6961 cgtttgcgcc tagcttcctg ctgaacatca aaggcaagaa aacatctgtt gtcaaagaca
     7021 gcatccttga acaaggacaa ttaacagtta acaaataaaa acgcaaaaga aaatgccgat
     7081 atcctattgg cattgacgtc aggtggcact tttcgcatgc cctggtcacc gtctcctcgg
     7141 cctccaccaa gggtccatcg gtcttccccc tggcaccctc ctccaagagc acctctgggg
     7201 gcacagcggc cctgggctgc ctggtcaagg actacttccc cgaaccggtg acggtgtcgt
     7261 ggaactcagg cgccctgacc agcggcgtgc acaccttccc ggctgtccta cagtcctcag
     7321 gactctactc cctcagcagc gtggtgaccg tgccctccag cagcttgggc acccagacct
     7381 acatctgcaa cgtgaatcac aagcccagca acaccaaggt cgacaagaaa gttgagccca
     7441 aatcttgtga caaaactcac acaggaggtt ctggcagcgc cggtggcctg aacgatattt
     7501 ttgaagcgca gaaaattgaa tggcatgaat gatgacgaag ctgtcgacaa aaaggtggaa
     7561 cctaagagct gcgataagac ccatacctaa taaggtctca tcgacaaaaa ggtcgaaccg
     7621 aagtcctgcg acaaaacaca cacgggtggc tctcatcatc accatcacca ctaataagct
     7681 tatcgctgat aagctgtcaa acatgagcag atctgagccc gcctaatgag cgggcttttt
     7741 tttcagatct gcttgaagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt
     7801 catgataata atggtttctt aggtgctcgt caaa
//
