LOCUS       sequence-459868-        7414 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 7414)
  AUTHORS   Zhu H, Kavsak P, Abdollah S, Wrana JL, Thomsen GH
  TITLE     A SMAD ubiquitin ligase targets the BMP pathway and affects 
            embryonic pattern formation.
  JOURNAL   Nature. 1999 Aug 12. 400(6745):687-93.
  PUBMED    10458166
REFERENCE   2  (bases 1 to 7414)
  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. 
            1999 Aug 12. 400(6745):687-93."
COMMENT     Sequence Label: pGEX4T1-Smurf1 wt
FEATURES             Location/Qualifiers
     source          1..7414
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(81..86,87..102,103..109)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    117..133
                     /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)."
     CDS             156..809
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     767..789
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             816..833
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             855..875
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(3176..3193)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     primer_bind     complement(3362..3384)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3529..3547)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3615..3719
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3720..3788,3789..4580)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3938..3957)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4751..5339
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5240..5259
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5493..5510
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        5583..5660
                     /gene="lacI"
                     /label=lacI promoter
     CDS             5661..6743
                     /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(5680..5699)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(6792..6797,6798..6815,6816..6822)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6830..6846
                     /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     6854..6870
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6854..6870
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             6866..7393
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(6886..6903)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6886..6902)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6895..6917)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 ctcaaggcgc actcccgttc tggataatgt tttttgcgcc gacatcataa cggttctggc
       61 aaatattctg aaatgagctg ttgacaatta atcatcggct cgtataatgt gtggaattgt
      121 gagcggataa caatttcaca caggaaacag tattcatgtc ccctatacta ggttattgga
      181 aaattaaggg ccttgtgcaa cccactcgac ttcttttgga atatcttgaa gaaaaatatg
      241 aagagcattt gtatgagcgc gatgaaggtg ataaatggcg aaacaaaaag tttgaattgg
      301 gtttggagtt tcccaatctt ccttattata ttgatggtga tgttaaatta acacagtcta
      361 tggccatcat acgttatata gctgacaagc acaacatgtt gggtggttgt ccaaaagagc
      421 gtgcagagat ttcaatgctt gaaggagcgg ttttggatat tagatacggt gtttcgagaa
      481 ttgcatatag taaagacttt gaaactctca aagttgattt tcttagcaag ctacctgaaa
      541 tgctgaaaat gttcgaagat cgtttatgtc ataaaacata tttaaatggt gatcatgtaa
      601 cccatcctga cttcatgttg tatgacgctc ttgatgttgt tttatacatg gacccaatgt
      661 gcctggatgc gttcccaaaa ttagtttgtt ttaaaaaacg tattgaagct atcccacaaa
      721 ttgataagta cttgaaatcc agcaagtata tagcatggcc tttgcagggc tggcaagcca
      781 cgtttggtgg tggcgaccat cctccaaaat cggatctggt tccgcgtgga tcggattacg
      841 atatcccaac gaccgaaaac ctgtattttc agggcgccat ggatccggaa ttcaaaggcc
      901 tacgtcgaca aagcaacgtg gtgacacgga gaggaggctc cagcatcaag atccgtctga
      961 cagtgttatg tgccaagaac cttgcaaaga aagacttctt caggctccct gacccttttg
     1021 caaagattgt cgtggatggg tctgggcagt gccactcaac cgacactgtg aaaaacacat
     1081 tggacccaaa gtggaaccag cactatgatc tatatgttgg gaaaacggat tcgataacca
     1141 ttagcgtgtg gaaccataag aaaattcaca agaaacaggg agctggcttc ctgggctgtg
     1201 tgcggctgct ctccaatgcc atcagcagat taaaagatac cggataccag cgtttggatc
     1261 tatgcaaact aaacccctca gatactgatg cagttcgtgg ccagatagtg gtcagtttac
     1321 agacacgaga cagaatagga accggcggct cggtggtgga ctgcagagga ctgttagaaa
     1381 atgaaggaac ggtgtatgaa gactccgggc ctgggaggcc gctcagctgc ttcatggagg
     1441 aaccagcccc ttacacagat agcaccggtg ctgctgctgg aggagggaat tgcaggttcg
     1501 tggagtcccc aagtcaagat caaagacttc aggcacagcg gcttcgaaac cctgatgtgc
     1561 gaggttcact acagacgccc cagaaccgac cacacggcca ccagtccccg gaactgcccg
     1621 aaggctacga acaaagaaca acagtccagg gccaagttta ctttttgcat acacagactg
     1681 gagttagcac gtggcacgac cccaggatac caagagacct taacagtgtg aactgtgatg
     1741 aacttggacc actgccgcca ggctgggaag tcagaagtac agtttctggg aggatatatt
     1801 ttgtagatca taataaccga acaacccagt ttacagaccc aaggttacac cacatcatga
     1861 atcaccagtg ccaactcaag gagcccagcc agccgctgcc actgcccagt gagggctctc
     1921 tggaggacga ggagcttcct gcccagagat acgaaagaga tctagtccag aagctgaaag
     1981 tcctcagaca cgaactgtcg cttcagcagc cccaagctgg tcattgccgc atcgaagtgt
     2041 ccagagaaga aatctttgag gagtcttacc gccagataat gaagatgcga ccgaaagact
     2101 tgaaaaaacg gctgatggtg aaattccgtg gggaagaagg tttggattac ggtggtgtgg
     2161 ccagggagtg gctttacttg ctgtgccatg aaatgctgaa tccttattac gggctcttcc
     2221 agtattctac ggacaatatt tacatgttgc aaataaatcc ggattcttca atcaaccccg
     2281 accacttgtc ttatttccac tttgtggggc ggatcatggg gctggctgtg ttccatggac
     2341 actacatcaa cgggggcttc acagtgccct tctacaagca gctgctgggg aagcccatcc
     2401 agctctcaga tctggaatct gtggacccag agctgcataa gagcttggtg tggatcctag
     2461 agaacgacat cacgcctgta ctggaccaca ccttctgcgt ggaacacaac gccttcgggc
     2521 ggatcctgca gcatgaactg aaacccaatg gcagaaatgt gccagtcaca gaggagaata
     2581 agaaagaata cgtccggttg tatgtaaact ggaggtttat gagaggaatc gaagcccagt
     2641 tcttagctct gcagaagggg ttcaatgagc tcatccctca acatctgctg aagccttttg
     2701 accagaagga actggagctg atcataggcg gcctggataa aatagacttg aacgactgga
     2761 agtcgaacac gcggctgaag cactgtgtgg ccgacagcaa catcgtgcgg tggttctggc
     2821 aagcggtgga gacgttcgat gaagaaagga gggccaggct cctgcagttt gtgactgggt
     2881 ccacgcgagt cccgctccaa ggcttcaagg ctttgcaagg ttctacaggc gcggcagggc
     2941 cccggctgtt caccatccac ctgatagacg cgaacacaga caaccttccg aaggcccata
     3001 cctgctttaa ccggatcgac attccaccat atgagtccta tgagaagctc tacgagaagc
     3061 tgctgacagc cgtggaggag acctgcgggt ttgctgtgga gtaatctaga gcctgcagtc
     3121 tcgaggcatg cggtaccaag cttggctgtt ttggcggatg agagaagatt ttcagcctga
     3181 tacagattaa atcagaacgc agaagcggtc tgataaaaca gaatttgcct ggcggcagta
     3241 gcgcggtggt cccacctgac cccatgccga actcagaagt gaaacgccgt agcgccgatg
     3301 gtacaattca tcgtgactga ctgacgatct gcctcgcgcg tttcggtgat gacggtgaaa
     3361 acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga
     3421 gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc gcagccatga
     3481 cccagtcacg tagcgatagc ggagtgtata attcttgaag acgaaagggc ctcgtgatac
     3541 gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt
     3601 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     3661 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     3721 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     3781 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     3841 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     3901 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     3961 gtgttgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     4021 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     4081 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     4141 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     4201 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     4261 ctgcagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     4321 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     4381 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     4441 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     4501 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     4561 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     4621 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     4681 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     4741 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     4801 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     4861 taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag
     4921 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     4981 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     5041 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     5101 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     5161 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     5221 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     5281 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     5341 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     5401 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg
     5461 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
     5521 agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcataaatt
     5581 ccgacaccat cgaatggcgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga
     5641 gtcaattcag ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg
     5701 gtgtctctta tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa
     5761 cgcgggaaaa agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac
     5821 aacaactggc gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc
     5881 acgcgccgtc gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg
     5941 tggtggtgtc gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc
     6001 ttctcgcgca acgcgtcagt gggctgatca ttaactatcc gctggatgac caggatgcca
     6061 ttgctgtgga agctgcctgc actaatgttc cggcgttatt tcttgatgtc tctgaccaga
     6121 cacccatcaa cagtattatt ttctcccatg aagacggtac gcgactgggc gtggagcatc
     6181 tggtcgcatt gggtcaccag caaatcgcgc tgttagcggg cccattaagt tctgtctcgg
     6241 cgcgtctgcg tctggctggc tggcataaat atctcactcg caatcaaatt cagccgatag
     6301 cggaacggga aggcgactgg agtgccatgt ccggttttca acaaaccatg caaatgctga
     6361 atgagggcat cgttcccact gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa
     6421 tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga tatctcggta gtgggatacg
     6481 acgataccga agacagctca tgttatatcc cgccgttaac caccatcaaa caggattttc
     6541 gcctgctggg gcaaaccagc gtggaccgct tgctgcaact ctctcagggc caggcggtga
     6601 agggcaatca gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata
     6661 cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt
     6721 cccgactgga aagcgggcag tgagcgcaac gcaattaatg taagttagct cactcattag
     6781 gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga
     6841 taacaatttc acacaggaaa cagctatgac catgattacg gattcactgg ccgtcgtttt
     6901 acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc
     6961 ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt
     7021 gcgcagcctg aatggcgaat ggcgctttgc ctggtttccg gcaccagaag cggtgccgga
     7081 aagctggctg gagtgcgatc ttcctgaggc cgatactgtc gtcgtcccct caaactggca
     7141 gatgcacggt tacgatgcgc ccatctacac caacgtgacc tatcccatta cggtcaatcc
     7201 gccgtttgtt cccacggaga atccgacggg ttgttactcg ctcacattta atgttgatga
     7261 aagctggcta caggaaggcc agacgcgaat tatttttgat ggcgttggaa ttagcttatc
     7321 gactgcacgg tgcaccaatg cttctggcgt caggcagcca tcggaagctg tggtatggct
     7381 gtgcaggtcg taaatcactg cataattcgt gtcg
//
