LOCUS       sequence-242767-        6815 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6815)
  AUTHORS   Tzivion G, Luo Z, Avruch J
  TITLE     A dimeric 14-3-3 protein is an essential cofactor for Raf kinase 
            activity.
  JOURNAL   Nature 1998 Jul 2;394(6688):88-92.
  PUBMED    9665134
REFERENCE   2  (bases 1 to 6815)
  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: "Nature"; 
            date: "1998-07-2"; volume: "394(6688)"; pages: "88-92"
COMMENT     Sequence Label: GST-14-3-3wt (pEBG)
FEATURES             Location/Qualifiers
     source          1..6815
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    112..133
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(148..153,154..171,172..178)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    186..202
                     /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     191..213
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     210..226
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     210..226
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        258..468
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        280..476
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      327..462
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        563..1741
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          794..1732
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1689..1709
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             1793..2446
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MAPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2404..2426
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2453..2470
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(3949..3966)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3949..3965)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3958..3980)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4178..4633
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4265..4284)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4475..4496
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4650..4669)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4769..4791
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4829..4847)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4915..5019
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5020..5088,5089..5880)
                     /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     complement(5238..5257)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6051..6639
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6540..6559
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6793..6810
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag
       61 ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag
      121 ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg
      181 tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa
      241 gcttaattcc ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
      301 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
      361 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct
      421 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc
      481 tttgcaaaga tggataaagt tttaaacaga gaggaatctt tgcagctaat ggaccttcta
      541 ggtcttgaaa ggagtgggaa ttggctccgg tgcccgtcag tgggcagagc gcacatcgcc
      601 cacagtcccc gagaagttgg ggggaggggt cggcaattga accggtgcct agagaaggtg
      661 gcgcggggta aactgggaaa gtgatgtcgt gtactggctc cgcctttttc ccgagggtgg
      721 gggagaaccg tatataagtg cagtagtcgc cgtgaacgtt ctttttcgca acgggtttgc
      781 cgccagaaca caggtaagtg ccgtgtgtgg ttcccgcggg cctggcctct ttacgggtta
      841 tggcccttgc gtgccttgaa ttacttccac ctggctgcag tacgtgattc ttgatcccga
      901 gcttcgggtt ggaagtgggt gggagagttc gaggccttgc gcttaaggag ccccttcgcc
      961 tcgtgcttga gttgaggcct ggcctgggcg ctggggccgc cgcgtgcgaa tctggtggca
     1021 ccttcgcgcc tgtctcgctg ctttcgataa gtctctagcc atttaaaatt tttgatgacc
     1081 tgctgcgacg ctttttttct ggcaagatag tcttgtaaat gcgggccaag atctgcacac
     1141 tggtatttcg gtttttgggg ccgcgggcgg cgacggggcc cgtgcgtccc agcgcacatg
     1201 ttcggcgagg cggggcctgc gagcgcggcc accgagaatc ggacgggggt agtctcaagc
     1261 tggccggcct gctctggtgc ctggcctcgc gccgccgtgt atcgccccgc cctgggcggc
     1321 aaggctggcc cggtcggcac cagttgcgtg agcggaaaga tggccgcttc ccggccctgc
     1381 tgcagggagc tcaaaatgga ggacgcggcg ctcgggagag cgggcgggtg agtcacccac
     1441 acaaaggaaa agggcctttc cgtcctcagc cgtcgcttca tgtgactcca cggagtaccg
     1501 ggcgccgtcc aggcacctcg attagttctc gagcttttgg agtacgtcgt ctttaggttg
     1561 gggggagggg ttttatgcga tggagtttcc ccacactgag tgggtggaga ctgaagttag
     1621 gccagcttgg cacttgatgt aattctcctt ggaatttgcc ctttttgagt ttggatcttg
     1681 gttcattctc aagcctcaga cagtggttca aagttttttt cttccatttc aggtgtcgtg
     1741 aggaattctc tagagatccc tcgacctcga gatccattgt gctggatcta caatggcccc
     1801 tatactaggt tattggaaaa ttaagggcct tgtgcaaccc actcgacttc ttttggaata
     1861 tcttgaagaa aaatatgaag agcatttgta tgagcgcgat gaaggtgata aatggcgaaa
     1921 caaaaagttt gaattgggtt tggagtttcc caatcttcct tattatattg atggtgatgt
     1981 taaattaaca cagtctatgg ccatcatacg ttatatagct gacaagcaca acatgttggg
     2041 tggttgtcca aaagagcgtg cagagatttc aatgcttgaa ggagcggttt tggatattag
     2101 atacggtgtt tcgagaattg catatagtaa agactttgaa actctcaaag ttgattttct
     2161 tagcaagcta cctgaaatgc tgaaaatgtt cgaagatcgt ttatgtcata aaacatattt
     2221 aaatggtgat catgtaaccc atcctgactt catgttgtat gacgctcttg atgttgtttt
     2281 atacatggac ccaatgtgcc tggatgcgtt cccaaaatta gtttgtttta aaaaacgtat
     2341 tgaagctatc ccacaaattg ataagtactt gaaatccagc aagtatatag catggccttt
     2401 gcagggctgg caagccacgt ttggtggtgg cgaccatcct ccaaaatcgg atctggttcc
     2461 gcgtggatcc ccgggaattt ccggtggtgg tggtggaatt caaatggata aaaatgagct
     2521 ggtgcagaag gccaagctgg ccgagcaggc agagcgatat gatgacatgg cagcctgcat
     2581 gaagtctgtc actgagcagg gagctgagct gtcgaatgag gagagaaacc ttctctctgt
     2641 tgcttataaa aacgttgtag gagcccgtag gtcatcgtgg agggtcgtct caagtattga
     2701 gcagaagacg gaaggtgctg agaaaaagca gcagatggct cgagaataca gagagaagat
     2761 cgagacggag ctgcgtgaca tctgcaacga tgtactgtct cttttggaaa agttcttgat
     2821 ccccaatgct tcgcaaccag aaagcaaagt cttctatttg aaaatgaagg gtgactacta
     2881 ccgttacttg gccgaggttg ctgctggtga tgacaagaaa ggaattgtgg accagtcaca
     2941 gcaagcatac caagaagcat ttgaaatcag caaaaaggag atgcagccga cacaccccat
     3001 cagactgggt ctggccctca acttctctgt gttctattac gagatcctga actccccaga
     3061 gaaagcctgc tctcttgcaa aaacagcttt cgatgaagcc attgctgaac ttgatacatt
     3121 aagtgaagag tcgtacaaag acagcacgct aataatgcag ttactgagag acaacttaac
     3181 attgtggaca tcggataccc aaggagatga agcagaagca ggagaaggag gggaaaatta
     3241 aggatccact agtggtacca tcgatgcggc cgcgactcta gagtgagggt ccccacctgg
     3301 gacccttgag agtatcaggt ctcccacgtg ggagacaaga aatccctgtt taatatttaa
     3361 acagcagtgt tccccatctg ggtccttgca cccctcactc tggcctcagc cgactgcaca
     3421 gcggcccctg catccccttg gctgtgaggc ccctggacaa gcagaggtgg ccagagctgg
     3481 gaggcatggc cctggggtcc cacgaatttg ctggggaatc tcgtttttct tcttaagact
     3541 tttgggacat ggtttgactc ccgaacatca ccgacgtgtc tcctgttttt ctgggtggcc
     3601 tcgggacacc tgccctgccc ccacgagggt caggactgtg actcttttta gggccaggca
     3661 ggtgcctgga catttgcctt gctggacggg gactggggat gtgggaggga gcagacagga
     3721 ggaatcatgt caggcctgtg tgtgaaagga agctccactg tcaccctcca cctcttcacc
     3781 ccccactcac cagtgtcccc tccactgtca cattgtaact gaacttcagg ataataaagw
     3841 gwttgcctcc aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa
     3901 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa ccgaattcac tggccgtcgt
     3961 tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca
     4021 tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca
     4081 gttgcgcagc ctgaatggcg aatggcgcct gatgcggtat tttctcctta cgcatctgtg
     4141 cggtatttca caccgcatac gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta
     4201 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccttagcg
     4261 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
     4321 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     4381 aaaaaacttg atttgggtga tggttcacgt agtgggccat cgccctgata gacggttttt
     4441 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca
     4501 acactcaact ctatctcggg ctattctttt gatttataag ggattttgcc gatttcggtc
     4561 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta
     4621 acgtttacaa ttttatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc
     4681 cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca
     4741 tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg
     4801 tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac gcctattttt ataggttaat
     4861 gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga
     4921 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     4981 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     5041 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     5101 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     5161 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     5221 agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag
     5281 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     5341 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     5401 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     5461 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     5521 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg
     5581 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     5641 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     5701 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     5761 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     5821 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     5881 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     5941 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     6001 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     6061 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     6121 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     6181 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     6241 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     6301 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     6361 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     6421 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     6481 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     6541 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     6601 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     6661 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     6721 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     6781 acgaccgagc gcagcgagtc agtgagcgag gaagc
//
