LOCUS       sequence-298394-        7739 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 7739)
  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 7739)
  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..7739
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        3..80
                     /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             81..1163
                     /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(100..119)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1406..1411,1412..1427,1428..1434)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1442..1458
                     /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     1447..1469
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1528..2628
                     /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="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     regulatory      2485..2494
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     2602..2625
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             2677..2688
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             2731..2751
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             3787..3804
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3831..3848)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3831..3847)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3840..3862)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(4026..4043)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      4196..4282
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4374..4401
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        4421..4512
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4513..4581,4582..5373)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERSPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4731..4750)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(5415..5928)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5610..5631)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5822..5841
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      6039..6627
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6528..6547
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6781..6798
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    6813..6953
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(6879..6898)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7039..7061
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(7055..7246)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 ccgacaccat cgaatggtgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga
       61 gtcaattcag ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg
      121 gtgtctctta tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa
      181 cgcgggaaaa agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac
      241 aacaactggc gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc
      301 acgcgccgtc gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg
      361 tggtggtgtc gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc
      421 ttctcgcgca acgcgtcagt gggctgatca ttaactatcc gctggatgac caggatgcca
      481 ttgctgtgga agctgcctgc actaatgttc cggcgttatt tcttgatgtc tctgaccaga
      541 cacccatcaa cagtattatt ttctcccatg aagacggtac gcgactgggc gtggagcatc
      601 tggtcgcatt gggtcaccag caaatcgcgc tgttagcggg cccattaagt tctgtctcgg
      661 cgcgtctgcg tctggctggc tggcataaat atctcactcg caatcaaatt cagccgatag
      721 cggaacggga aggcgactgg agtgccatgt ccggttttca acaaaccatg caaatgctga
      781 atgagggcat cgttcccact gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa
      841 tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga tatctcggta gtgggatacg
      901 acgataccga agacagctca tgttatatcc cgccgttaac caccatcaaa caggattttc
      961 gcctgctggg gcaaaccagc gtggaccgct tgctgcaact ctctcagggc caggcggtga
     1021 agggcaatca gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata
     1081 cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt
     1141 cccgactgga aagcgggcag tgagcgcaac gcaattaatg taagttagct cactcattag
     1201 gcacaattct catgtttgac agcttatcat cgactgcacg gtgcaccaat gcttctggcg
     1261 tcaggcagcc atcggaagct gtggtatggc tgtgcaggtc gtaaatcact gcataattcg
     1321 tgtcgctcaa ggcgcactcc cgttctggat aatgtttttt gcgccgacat cataacggtt
     1381 ctggcaaata ttctgaaatg agctgttgac aattaatcat cggctcgtat aatgtgtgga
     1441 attgtgagcg gataacaatt tcacacagga aacagccagt ccgtttaggt gttttcacga
     1501 gcacttcacc aacaaggacc atagattatg aaaatcgaag aaggtaaact ggtaatctgg
     1561 attaacggcg ataaaggcta taacggtctc gctgaagtcg gtaagaaatt cgagaaagat
     1621 accggaatta aagtcaccgt tgagcatccg gataaactgg aagagaaatt cccacaggtt
     1681 gcggcaactg gcgatggccc tgacattatc ttctgggcac acgaccgctt tggtggctac
     1741 gctcaatctg gcctgttggc tgaaatcacc ccggacaaag cgttccagga caagctgtat
     1801 ccgtttacct gggatgccgt acgttacaac ggcaagctga ttgcttaccc gatcgctgtt
     1861 gaagcgttat cgctgattta taacaaagat ctgctgccga acccgccaaa aacctgggaa
     1921 gagatcccgg cgctggataa agaactgaaa gcgaaaggta agagcgcgct gatgttcaac
     1981 ctgcaagaac cgtacttcac ctggccgctg attgctgctg acgggggtta tgcgttcaag
     2041 tatgaaaacg gcaagtacga cattaaagac gtgggcgtgg ataacgctgg cgcgaaagcg
     2101 ggtctgacct tcctggttga cctgattaaa aacaaacaca tgaatgcaga caccgattac
     2161 tccatcgcag aagctgcctt taataaaggc gaaacagcga tgaccatcaa cggcccgtgg
     2221 gcatggtcca acatcgacac cagcaaagtg aattatggtg taacggtact gccgaccttc
     2281 aagggtcaac catccaaacc gttcgttggc gtgctgagcg caggtattaa cgccgccagt
     2341 ccgaacaaag agctggcaaa agagttcctc gaaaactatc tgctgactga tgaaggtctg
     2401 gaagcggtta ataaagacaa accgctgggt gccgtagcgc tgaagtctta cgaggaagag
     2461 ttggcgaaag atccacgtat tgccgccacc atggaaaacg cccagaaagg tgaaatcatg
     2521 ccgaacatcc cgcagatgtc cgctttctgg tatgccgtgc gtactgcggt gatcaacgcc
     2581 gccagcggtc gtcagactgt cgatgaagcc ctgaaagacg cgcagactaa ttcgagctcg
     2641 aacaacaaca acaataacaa taacaacaac ctcgggatcg agggaaggat ttcaactagt
     2701 ggatctggtg gtggtggcgg ttccatgagc gagaatcttt attttcaggg atccatgacc
     2761 atggaatctg gagccgagaa ccagcagagt ggagatgcag ctgtaacaga agctgaaaac
     2821 caacaaatga cagttcaagc ccagccacag attgccacat tagcccaggt atctatgcca
     2881 gcagctcatg caacatcatc tgctcccacc gtaactctag tacagctgcc caatgggcag
     2941 acagttcaag tccatggagt cattcaggcg gcccagccat cagttattca gtctccacaa
     3001 gtccaaacag ttcagtcttc ctgtaaggac ttaaaaagac ttttctccgg aacacagatt
     3061 tcaactattg cagaaagtga agattcacag gagtcagtgg atagtgtaac tgattcccaa
     3121 aagcgaaggg aaattctttc aaggaggcct tcctacagga aaattttgaa tgacttatct
     3181 tctgatgcac caggagtgcc aaggattgaa gaagagaagt ctgaagagga gacttcagca
     3241 cctgccatca ccactgtaac ggtgccaact ccaatttacc aaactagcag tggacagtat
     3301 attgccatta cccagggagg agcaatacag ctggctaaca atggtaccga tggggtacag
     3361 ggcctgcaaa cattaaccat gaccaatgca gcagccactc agccgggtac taccattcta
     3421 cagtatgcac agaccactga tggacagcag atcttagtgc ccagcaacca agttgttgtt
     3481 caagctgcct ctggagacgt acaaacatac cagattcgca cagcacccac tagcactatt
     3541 gcccctggag ttgttatggc atcctcccca gcacttccta cacagcctgc tgaagaagca
     3601 gcacgaaaga gagaggtccg tctaatgaag aacagggaag cagctcgaga gtgtcgtaga
     3661 aagaagaaag aatatgtgaa atgtttagaa aacagagtgg cagtgcttga aaatcaaaac
     3721 aagacattga ttgaggagct aaaagcactt aaggaccttt actgccacaa atcagatagc
     3781 ggccgccacc atcaccacca ccactagggg ccgcactcga gaagcttggc actggccgtc
     3841 gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca
     3901 catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa
     3961 cagttgcgca gcctgaatgg cgaatggcag cttggctgtt ttggcggatg agataagatt
     4021 ttcagcctga tacagattaa atcagaacgc agaagcggtc tgataaaaca gaatttgcct
     4081 ggcggcagta gcgcggtggt cccacctgac cccatgccga actcagaagt gaaacgccgt
     4141 agcgccgatg gtagtgtggg gtctccccat gcgagagtag ggaactgcca ggcatcaaat
     4201 aaaacgaaag gctcagtcga aagactgggc ctttcgtttt atctgttgtt tgtcggtgaa
     4261 cgctctcctg agtaggacaa atccgccggg agcggatttg aacgttgcga agcaacggcc
     4321 cggagggtgg cgggcaggac gcccgccata aactgccagg catcaaatta agcagaaggc
     4381 catcctgacg gatggccttt ttgcgtttct acaaactctt tttgtttatt tttctaaata
     4441 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     4501 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     4561 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     4621 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     4681 agttttcgcc ccgaagaacg ttctccaatg atgagcactt ttaaagttct gctatgtggc
     4741 gcggtattat cccgtgttga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     4801 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     4861 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     4921 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     4981 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     5041 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     5101 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     5161 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     5221 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     5281 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     5341 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     5401 ctttagattg atttaccccg gttgataatc agaaaagccc caaaaacagg aagattgtat
     5461 aagcaaatat ttaaattgta aacgttaata ttttgttaaa attcgcgtta aatttttgtt
     5521 aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag
     5581 aatagcccga gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga
     5641 acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg
     5701 aaccatcacc caaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc
     5761 ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg
     5821 aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc
     5881 gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtaa aaggatctag
     5941 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     6001 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     6061 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     6121 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     6181 actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     6241 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     6301 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     6361 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     6421 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     6481 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     6541 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     6601 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     6661 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     6721 aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc
     6781 agcgagtcag tgagcgagga agcggaagag cgcctgatgc ggtattttct ccttacgcat
     6841 ctgtgcggta tttcacaccg catatggtgc actctcagta caatctgctc tgatgccgca
     6901 tagttaagcc agtatacact ccgctatcgc tacgtgactg ggtcatggct gcgccccgac
     6961 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
     7021 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
     7081 aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc gtgcagcgat tcacagatgt
     7141 ctgcctgttc atccgcgtcc agctcgttga gtttctccag aagcgttaat gtctggcttc
     7201 tgataaagcg ggccatgtta agggcggttt tttcctgttt ggtcactgat gcctccgtgt
     7261 aagggggatt tctgttcatg ggggtaatga taccgatgaa acgagagagg atgctcacga
     7321 tacgggttac tgatgatgaa catgcccggt tactggaacg ttgtgagggt aaacaactgg
     7381 cggtatggat gcggcgggac cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta
     7441 atacagatgt aggtgttcca cagggtagcc agcagcatcc tgcgatgcag atccggaaca
     7501 taatggtgca gggcgctgac ttccgcgttt ccagacttta cgaaacacgg aaaccgaaga
     7561 ccattcatgt tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct
     7621 cgcgtatcgg tgattcattc tgctaaccag taaggcaacc ccgccagcct agccgggtcc
     7681 tcaacgacag gagcacgatc atgcgcaccc gtggccagga cccaacgctg cccgaaatt
//
