LOCUS       sequence-308814-        7738 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses MBP-CREB-His (human CREB as a fusion protein with MBP and 
            His- tags) in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7738)
  AUTHORS   Gavagan M, Fagnan E, Speltz EB, Zalatan JG
  TITLE     The Scaffold Protein Axin Promotes Signaling Specificity within the 
            Wnt Pathway by Suppressing Competing Kinase Reactions.
  JOURNAL   Cell Syst. 2020 Jun 24;10(6):515-525.e5. doi: 
            10.1016/j.cels.2020.05.002. Epub 2020 Jun 17.
  PUBMED    32553184
REFERENCE   2  (bases 1 to 7738)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.cels.2020.05.002"; journalName: "Cell Syst"; date: 
            "2020-06-24- 24"; volume: "10"; issue: "6"; pages: "515-525.e5"
COMMENT     Sequence Label: pMG059 MBP-CREB-His
FEATURES             Location/Qualifiers
     source          1..7738
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..78
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             79..1161
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(98..117)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1404..1409,1410..1425,1426..1432)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1440..1456
                     /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)."
     primer_bind     1445..1467
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1526..2626
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKTEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     regulatory      2483..2492
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     2600..2623
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             2675..2686
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             2729..2749
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             3785..3802
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3829..3846)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3829..3845)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3838..3860)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(4024..4041)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      4194..4280
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4372..4399
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        4418..4509
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4510..4578,4579..5370)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4728..4747)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(5412..5925)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5607..5628)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5819..5838
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      6036..6624
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6525..6544
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6778..6795
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    6810..6950
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(6876..6895)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7036..7058
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(7052..7243)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 gacaccatcg aatggtgcaa aacctttcgc ggtatggcat gatagcgccc ggaagagagt
       61 caattcaggg tggtgaatgt gaaaccagta acgttatacg atgtcgcaga gtatgccggt
      121 gtctcttatc agaccgtttc ccgcgtggtg aaccaggcca gccacgtttc tgcgaaaacg
      181 cgggaaaaag tggaagcggc gatggcggag ctgaattaca ttcccaaccg cgtggcacaa
      241 caactggcgg gcaaacagtc gttgctgatt ggcgttgcca cctccagtct ggccctgcac
      301 gcgccgtcgc aaattgtcgc ggcgattaaa tctcgcgccg atcaactggg tgccagcgtg
      361 gtggtgtcga tggtagaacg aagcggcgtc gaagcctgta aagcggcggt gcacaatctt
      421 ctcgcgcaac gcgtcagtgg gctgatcatt aactatccgc tggatgacca ggatgccatt
      481 gctgtggaag ctgcctgcac taatgttccg gcgttatttc ttgatgtctc tgaccagaca
      541 cccatcaaca gtattatttt ctcccatgaa gacggtacgc gactgggcgt ggagcatctg
      601 gtcgcattgg gtcaccagca aatcgcgctg ttagcgggcc cattaagttc tgtctcggcg
      661 cgtctgcgtc tggctggctg gcataaatat ctcactcgca atcaaattca gccgatagcg
      721 gaacgggaag gcgactggag tgccatgtcc ggttttcaac aaaccatgca aatgctgaat
      781 gagggcatcg ttcccactgc gatgctggtt gccaacgatc agatggcgct gggcgcaatg
      841 cgcgccatta ccgagtccgg gctgcgcgtt ggtgcggata tctcggtagt gggatacgac
      901 gataccgaag acagctcatg ttatatcccg ccgttaacca ccatcaaaca ggattttcgc
      961 ctgctggggc aaaccagcgt ggaccgcttg ctgcaactct ctcagggcca ggcggtgaag
     1021 ggcaatcagc tgttgcccgt ctcactggtg aaaagaaaaa ccaccctggc gcccaatacg
     1081 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc
     1141 cgactggaaa gcgggcagtg agcgcaacgc aattaatgta agttagctca ctcattaggc
     1201 acaattctca tgtttgacag cttatcatcg actgcacggt gcaccaatgc ttctggcgtc
     1261 aggcagccat cggaagctgt ggtatggctg tgcaggtcgt aaatcactgc ataattcgtg
     1321 tcgctcaagg cgcactcccg ttctggataa tgttttttgc gccgacatca taacggttct
     1381 ggcaaatatt ctgaaatgag ctgttgacaa ttaatcatcg gctcgtataa tgtgtggaat
     1441 tgtgagcgga taacaatttc acacaggaaa cagccagtcc gtttaggtgt tttcacgagc
     1501 acttcaccaa caaggaccat agattatgaa aactgaagaa ggtaaactgg taatctggat
     1561 taacggcgat aaaggctata acggtctcgc tgaagtcggt aagaaattcg agaaagatac
     1621 cggaattaaa gtcaccgttg agcatccgga taaactggaa gagaaattcc cacaggttgc
     1681 ggcaactggc gatggccctg acattatctt ctgggcacac gaccgctttg gtggctacgc
     1741 tcaatctggc ctgttggctg aaatcacccc ggacaaagcg ttccaggaca agctgtatcc
     1801 gtttacctgg gatgccgtac gttacaacgg caagctgatt gcttacccga tcgctgttga
     1861 agcgttatcg ctgatttata acaaagatct gctgccgaac ccgccaaaaa cctgggaaga
     1921 gatcccggcg ctggataaag aactgaaagc gaaaggtaag agcgcgctga tgttcaacct
     1981 gcaagaaccg tacttcacct ggccgctgat tgctgctgac gggggttatg cgttcaagta
     2041 tgaaaacggc aagtacgaca ttaaagacgt gggcgtggat aacgctggcg cgaaagcggg
     2101 tctgaccttc ctggttgacc tgattaaaaa caaacacatg aatgcagaca ccgattactc
     2161 catcgcagaa gctgccttta ataaaggcga aacagcgatg accatcaacg gcccgtgggc
     2221 atggtccaac atcgacacca gcaaagtgaa ttatggtgta acggtactgc cgaccttcaa
     2281 gggtcaacca tccaaaccgt tcgttggcgt gctgagcgca ggtattaacg ccgccagtcc
     2341 gaacaaagag ctggcaaaag agttcctcga aaactatctg ctgactgatg aaggtctgga
     2401 agcggttaat aaagacaaac cgctgggtgc cgtagcgctg aagtcttacg aggaagagtt
     2461 ggcgaaagat ccacgtattg ccgccaccat ggaaaacgcc cagaaaggtg aaatcatgcc
     2521 gaacatcccg cagatgtccg ctttctggta tgccgtgcgt actgcggtga tcaacgccgc
     2581 cagcggtcgt cagactgtcg atgaagccct gaaagacgcg cagactaatt cgagctcgaa
     2641 caacaacaac aataacaata acaacaacct cgggatcgag ggaaggattt caactagtgg
     2701 atctggtggt ggtggcggtt ccatgagcga gaatctttat tttcagggat ccatgaccat
     2761 ggaatctgga gccgagaacc agcagagtgg agatgcagct gtaacagaag ctgaaaacca
     2821 acaaatgaca gttcaagccc agccacagat tgccacatta gcccaggtat ctatgccagc
     2881 agctcatgca acatcatctg ctcccaccgt aactctagta cagctgccca atgggcagac
     2941 agttcaagtc catggagtca ttcaggcggc ccagccatca gttattcagt ctccacaagt
     3001 ccaaacagtt cagtcttcct gtaaggactt aaaaagactt ttctccggaa cacagatttc
     3061 aactattgca gaaagtgaag attcacagga gtcagtggat agtgtaactg attcccaaaa
     3121 gcgaagggaa attctttcaa ggaggccttc ctacaggaaa attttgaatg acttatcttc
     3181 tgatgcacca ggagtgccaa ggattgaaga agagaagtct gaagaggaga cttcagcacc
     3241 tgccatcacc actgtaacgg tgccaactcc aatttaccaa actagcagtg gacagtatat
     3301 tgccattacc cagggaggag caatacagct ggctaacaat ggtaccgatg gggtacaggg
     3361 cctgcaaaca ttaaccatga ccaatgcagc agccactcag ccgggtacta ccattctaca
     3421 gtatgcacag accactgatg gacagcagat cttagtgccc agcaaccaag ttgttgttca
     3481 agctgcctct ggagacgtac aaacatacca gattcgcaca gcacccacta gcactattgc
     3541 ccctggagtt gttatggcat cctccccagc acttcctaca cagcctgctg aagaagcagc
     3601 acgaaagaga gaggtccgtc taatgaagaa cagggaagca gctcgagagt gtcgtagaaa
     3661 gaagaaagaa tatgtgaaat gtttagaaaa cagagtggca gtgcttgaaa atcaaaacaa
     3721 gacattgatt gaggagctaa aagcacttaa ggacctttac tgccacaaat cagatagcgg
     3781 ccgccaccat caccaccacc actaggggcc gcactcgaga agcttggcac tggccgtcgt
     3841 tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca
     3901 tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca
     3961 gttgcgcagc ctgaatggcg aatggcagct tggctgtttt ggcggatgag ataagatttt
     4021 cagcctgata cagattaaat cagaacgcag aagcggtctg ataaaacaga atttgcctgg
     4081 cggcagtagc gcggtggtcc cacctgaccc catgccgaac tcagaagtga aacgccgtag
     4141 cgccgatggt agtgtggggt ctccccatgc gagagtaggg aactgccagg catcaaataa
     4201 aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg tcggtgaacg
     4261 ctctcctgag taggacaaat ccgccgggag cggatttgaa cgttgcgaag caacggcccg
     4321 gagggtggcg ggcaggacgc ccgccataaa ctgccaggca tcaaattaag cagaaggcca
     4381 tcctgacgga tggccttttt gcgtttctac aaactctttt gtttattttt ctaaatacat
     4441 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     4501 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     4561 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     4621 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     4681 tttcgccccg aagaacgttt cccaatgatg agcactttta aagttctgct atgtggcgcg
     4741 gtattatccc gtgttgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     4801 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     4861 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     4921 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     4981 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     5041 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     5101 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     5161 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     5221 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     5281 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     5341 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     5401 tagattgatt taccccggtt gataatcaga aaagccccaa aaacaggaag attgtataag
     5461 caaatattta aattgtaaac gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa
     5521 tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat
     5581 agcccgagat agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg
     5641 tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac
     5701 catcacccaa atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta
     5761 aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag
     5821 ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg
     5881 taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtaaaag gatctaggtg
     5941 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     6001 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     6061 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     6121 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     6181 gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     6241 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     6301 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     6361 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     6421 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     6481 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     6541 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     6601 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     6661 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     6721 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     6781 gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt attttctcct tacgcatctg
     6841 tgcggtattt cacaccgcat atggtgcact ctcagtacaa tctgctctga tgccgcatag
     6901 ttaagccagt atacactccg ctatcgctac gtgactgggt catggctgcg ccccgacacc
     6961 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
     7021 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac
     7081 gcgcgaggca gctgcggtaa agctcatcag cgtggtcgtg cagcgattca cagatgtctg
     7141 cctgttcatc cgcgtccagc tcgttgagtt tctccagaag cgttaatgtc tggcttctga
     7201 taaagcgggc catgttaagg gcggtttttt cctgtttggt cactgatgcc tccgtgtaat
     7261 ggggatttct gttcatgggg gtaatgatac cgatgaaacg agagaggatg ctcacgatac
     7321 gggttactga tgatgaacat gcccggttac tggaacgttg tgagggtaaa caactggcgg
     7381 tatggatgcg gcgggaccag agaaaaatca ctcagggtca atgccagcgc ttcgttaata
     7441 cagatgtagg tgttccacag ggtagccagc agcatcctgc gatgcagatc cggaacataa
     7501 tggtgcaggg cgctgacttc cgcgtttcca gactttacga aacacggaaa ccgaagacca
     7561 ttcatgttgt tgctcaggtc gcagacgttt tgcagcagca gtcgcttcac gttcgctcgc
     7621 gtatcggtga ttcattctgc taaccagtaa ggcaaccccg ccagcctagc cgggtcctca
     7681 acgacaggag cacgatcatg cgcacccgtg gccaggaccc aacgctgccc gaaattcc
//
