LOCUS       sequence-244761-        6908 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of human GSK3 beta constitutively active mutant
            with HA tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6908)
  AUTHORS   Stambolic V, Woodgett JR
  TITLE     Mitogen inactivation of glycogen synthase kinase-3 beta in intact 
            cells via serine 9 phosphorylation.
  JOURNAL   Biochem J. 1994 Nov 1;303 ( Pt 3)(Pt 3):701-4. doi: 
            10.1042/bj3030701.
  PUBMED    7980435
REFERENCE   2  (bases 1 to 6908)
  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"; journalName: "Biochem 
            J."; date: "1994-11-1"; volume: "303 ( Pt 3)(Pt 3):701-4. doi"; 
            pages: " 10.1042/bj3030701"
COMMENT     Sequence Label: HA GSK3 beta S9A pcDNA3
FEATURES             Location/Qualifiers
     source          1..6908
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1787..1798
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     CDS             2233..2259
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2239..2265
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     promoter        complement(2461..2479)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2462..2479)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2499..2516)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2505..2729
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2775..3203
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2862..2881)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3072..3093
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3212..3232)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3217..3546
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3397..3532
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3459..3478
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3613..4407
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3667..3686)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4277..4296
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4581..4702
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4618..4637)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4672..4691
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4751..4767)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4751..4767)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4764..4786)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4775..4791
                     /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)."
     promoter        complement(join(4799..4805,4806..4823,4824..4829))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4844..4865
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4982..4999)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5153..5741)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5233..5252)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5912..6703,6704..6772))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6535..6554
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6773..6877)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttggtacc
      901 gagctcggat ccactagtaa cggccgccag tgtgctggaa ttctgcagat catgggaatt
      961 cccctcgaca tggcagggcg gcccagaacc accgcctttg cggagagctg caagccggtg
     1021 cagcagcctt cagcttttgg cagcatgaaa gttagcagag acaaggacgg cagcaaggtg
     1081 acaacagtgg tggcaactcc tgggcagggt ccagacaggc cacaagaagt cagctataca
     1141 gacactaaag tgattggaaa tggatcattt ggtgtggtat atcaagccaa actttgtgat
     1201 tcaggagaac tggtcgccat caagaaagta ttgcaggaca agagatttaa gaatcgagag
     1261 ctccagatca tgagaaagct agatcactgt aacatagtcc gattgcgtta tttcttctac
     1321 tccagtggtg agaagaaaga tgaggtctat cttaatctgg tgctggacta tgttccggaa
     1381 acagtataca gagttgccag acactatagt cgagccaaac agacgctccc tgtgatttat
     1441 gtcaagttgt atatgtatca gctgttccga agtttagcct atatccattc ctttggaatc
     1501 tgccatcggg atattaaacc gcagaacctc ttgttggatc ctgatactgc tgtattaaaa
     1561 ctctgtgact ttggaagtgc aaagcagctg gtccgaggag aacccaatgt ttcgtatatc
     1621 tgttctcggt actatagggc accagagttg atctttggag ccactgatta tacctctagt
     1681 atagatgtat ggtctgctgg ctgtgtgttg gctgagctgt tactaggaca accaatattt
     1741 ccaggggata gtggtgtgga tcagttggta gaaataatca aggtcctggg aactccaaca
     1801 agggagcaaa tcagagaaat gaacccaaac tacacagaat ttaaattccc tcaaattaag
     1861 gcacatcctt ggactaaggt cttccgaccc cgaactccac cggaggcaat tgcactgtgt
     1921 agccgtctgc tggagtatac accaactgcc cgactaacac cactggaagc ttgtgcacat
     1981 tcattttttg atgaattacg ggacccaaat gtcaaactac caaatgggcg agacacacct
     2041 gcactcttca acttcaccac tcaagaactg tcaagtaatc cacctctggc taccatcctt
     2101 attcctcctc atgctcggat tcaagcagct gcttcaaccc ccacaaatgc cacagcagcg
     2161 tcagatgcta atactggaga ccgtggacag accaataatg ctgcttctgc atcagcttcc
     2221 aactccacta gttacccata cgatgttcca gattacgcaa gcttgggtgg tcccaactag
     2281 tacagtcccg acggatcctc tagaggatcc ccgggcgagc tcccaaaaaa aaaaaaaaaa
     2341 aaaaaaaaaa aaaaccgaat tggggagctc actagtggat ccctccagct cgagaggcct
     2401 aattaattaa gtcatccatc acactggcgg ccgctcgagc atgcatctag agggccctat
     2461 tctatagtgt cacctaaatg ctagagctcg ctgatcagcc tcgactgtgc cttctagttg
     2521 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
     2581 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc
     2641 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag
     2701 gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc
     2761 tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     2821 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     2881 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     2941 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     3001 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     3061 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
     3121 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
     3181 ttaacaaaaa tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag
     3241 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     3301 aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     3361 tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt
     3421 tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc
     3481 gcctctgcct ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt
     3541 tgcaaaaagc tcccgggagc ttgtatatcc attttcggat ctgatcaaga gacaggatga
     3601 ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg
     3661 gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg
     3721 ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc
     3781 ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct
     3841 tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa
     3901 gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg
     3961 gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa
     4021 gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat
     4081 gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg
     4141 cgcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc
     4201 atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac
     4261 cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg
     4321 gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc
     4381 tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag
     4441 cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg
     4501 gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc
     4561 tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca
     4621 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     4681 ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg
     4741 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     4801 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     4861 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     4921 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     4981 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     5041 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     5101 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     5161 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     5221 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     5281 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     5341 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     5401 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     5461 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     5521 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     5581 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     5641 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     5701 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     5761 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     5821 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     5881 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     5941 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     6001 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     6061 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     6121 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     6181 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     6241 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     6301 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     6361 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     6421 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     6481 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     6541 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     6601 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     6661 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     6721 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     6781 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     6841 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     6901 ctgacgtc
//
