LOCUS       sequence-308824-        6814 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses MBP-GS 634-666-His (human 33aa Glycogen Synthase peptide 
            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 6814)
  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 6814)
  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: pMG058 MBP-GS 634-666-His
FEATURES             Location/Qualifiers
     source          1..6814
                     /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             2861..2878
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2905..2922)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2905..2921)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2914..2936)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(3100..3117)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      3270..3356
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3448..3475
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        3494..3585
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3586..3654,3655..4446)
                     /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(3804..3823)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(4488..5001)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4683..4704)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4895..4914
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      5112..5700
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5601..5620
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5854..5871
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5886..6026
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5952..5971)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6112..6134
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(6128..6319)
                     /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 cctaccgcta
     2761 cccacggcca gcctcggtgc caccgtcgcc ctcgctgtca cgacactcca gcccgcacca
     2821 gagtgaggac gaggaggatc cccggaacgg gagcggccgc caccatcacc accaccacta
     2881 ggggccgcac tcgagaagct tggcactggc cgtcgtttta caacgtcgtg actgggaaaa
     2941 ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa
     3001 tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg
     3061 gcagcttggc tgttttggcg gatgagataa gattttcagc ctgatacaga ttaaatcaga
     3121 acgcagaagc ggtctgataa aacagaattt gcctggcggc agtagcgcgg tggtcccacc
     3181 tgaccccatg ccgaactcag aagtgaaacg ccgtagcgcc gatggtagtg tggggtctcc
     3241 ccatgcgaga gtagggaact gccaggcatc aaataaaacg aaaggctcag tcgaaagact
     3301 gggcctttcg ttttatctgt tgtttgtcgg tgaacgctct cctgagtagg acaaatccgc
     3361 cgggagcgga tttgaacgtt gcgaagcaac ggcccggagg gtggcgggca ggacgcccgc
     3421 cataaactgc caggcatcaa attaagcaga aggccatcct gacggatggc ctttttgcgt
     3481 ttctacaaac tcttttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     3541 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
     3601 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     3661 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
     3721 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgtttccca
     3781 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtgt tgacgccggg
     3841 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
     3901 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     3961 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     4021 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     4081 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
     4141 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     4201 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     4261 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
     4321 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     4381 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     4441 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttacc ccggttgata
     4501 atcagaaaag ccccaaaaac aggaagattg tataagcaaa tatttaaatt gtaaacgtta
     4561 atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg
     4621 ccgaaatcgg caaaatccct tataaatcaa aagaatagcc cgagataggg ttgagtgttg
     4681 ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa
     4741 aaaccgtcta tcagggcgat ggcccactac gtgaaccatc acccaaatca agttttttgg
     4801 ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt
     4861 gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg
     4921 ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta
     4981 atgcgccgct acagggcgcg taaaaggatc taggtgaaga tcctttttga taatctcatg
     5041 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     5101 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     5161 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     5221 gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta
     5281 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     5341 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     5401 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     5461 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     5521 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     5581 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     5641 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     5701 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     5761 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
     5821 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
     5881 gagcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcatatgg
     5941 tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatac actccgctat
     6001 cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc aacacccgct gacgcgccct
     6061 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     6121 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct
     6181 catcagcgtg gtcgtgcagc gattcacaga tgtctgcctg ttcatccgcg tccagctcgt
     6241 tgagtttctc cagaagcgtt aatgtctggc ttctgataaa gcgggccatg ttaagggcgg
     6301 ttttttcctg tttggtcact gatgcctccg tgtaatgggg atttctgttc atgggggtaa
     6361 tgataccgat gaaacgagag aggatgctca cgatacgggt tactgatgat gaacatgccc
     6421 ggttactgga acgttgtgag ggtaaacaac tggcggtatg gatgcggcgg gaccagagaa
     6481 aaatcactca gggtcaatgc cagcgcttcg ttaatacaga tgtaggtgtt ccacagggta
     6541 gccagcagca tcctgcgatg cagatccgga acataatggt gcagggcgct gacttccgcg
     6601 tttccagact ttacgaaaca cggaaaccga agaccattca tgttgttgct caggtcgcag
     6661 acgttttgca gcagcagtcg cttcacgttc gctcgcgtat cggtgattca ttctgctaac
     6721 cagtaaggca accccgccag cctagccggg tcctcaacga caggagcacg atcatgcgca
     6781 cccgtggcca ggacccaacg ctgcccgaaa ttcc
//
