LOCUS       sequence-456656-        7280 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 7280)
  AUTHORS   Kavsak P, Rasmussen RK, Causing CG, Bonni S, Zhu H, Thomsen GH, 
            Wrana JL
  TITLE     Smad7 binds to Smurf2 to form an E3 ubiquitin ligase that targets 
            the TGF beta receptor for degradation.
  JOURNAL   Mol Cell. 2000 Dec . 6(6):1365-75.
  PUBMED    11163210
REFERENCE   2  (bases 1 to 7280)
  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: "Mol Cell. 
            2000 Dec . 6(6):1365-75."
COMMENT     Sequence Label: pGEX4T1-Smurf2 wt
FEATURES             Location/Qualifiers
     source          1..7280
                     /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(3228..3250)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3395..3413)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3481..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
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3804..3823)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4617..5205
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5106..5125
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5359..5376
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        5449..5526
                     /gene="lacI"
                     /label=lacI promoter
     CDS             5527..6609
                     /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(5546..5565)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(6658..6663,6664..6681,6682..6688)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6696..6712
                     /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     6720..6736
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6720..6736
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             6732..7259
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(6752..6769)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6752..6768)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6761..6783)
                     /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 agggcgccca tatgggagga tccccggaat
      901 tcccgggtcg acaatctaac cccggacgcc ggaggaacgg gcccgtcaag ctgcgcctga
      961 cagtactctg tgcaaaaaac ctggtgaaaa aggatttttt ccgacttcct gatccatttg
     1021 ctaaggtggt ggttgatgga tctgggcaat gccattctac agatactgtg aagaatacgc
     1081 ttgatccaaa gtggaatcag cattatgacc tgtatattgg aaagtctgat tcagttacga
     1141 tcagtgtatg gaatcacaag aagatccata agaaacaagg tgctggattt ctcggttgtg
     1201 ttcgtcttct ttccaatgcc atcaaccgcc tcaaagacac tggttatcag aggttggatt
     1261 tatgcaaact cgggccaaat gacaatgata cagttagagg acagatagta gtaagtcttc
     1321 agtccagaga ccgaataggc acaggaggac aagttgtgga ctgcagtcgt ttatttgata
     1381 acgatttacc agacggctgg gaagaaagga gaaccgcctc tggaagaatc cagtatctaa
     1441 accatataac aagaactacg caatgggagc gcccaacacg accggcatcc gaatattcta
     1501 gccctggcag acctcttagc tgctttgttg atgagaacac tccaattagt ggaacaaatg
     1561 gtgcaacatg tggacagtct tcagatccca ggctggcaga gaggagagtc aggtcacaac
     1621 gacatagaaa ttacatgagc agaacacatt tacatactcc tccagaccta ccagaaggct
     1681 atgaacagag gacaacgcaa caaggccagg tgtatttctt acatacacag actggtgtga
     1741 gcacatggca tgatccaaga gtgcccaggg atcttagcaa catcaattgt gaagagcttg
     1801 gtccattgcc tcctggatgg gagatccgta atacggcaac aggcagagtt tatttcgttg
     1861 accataacaa cagaacaaca caatttacgg atcctcggct gtctgctaac ttgcatttag
     1921 ttttaaatcg gcagaaccaa ttgaaagacc aacagcaaca gcaagtggta tcgttatgtc
     1981 ctgatgacac agaatgcctg acagtcccaa ggtacaagcg agacctggtt cagaaactaa
     2041 aaattttgcg gcaagaactt tcccaacaac agcctcaggc aggtcattgc cgcattgagg
     2101 tttccaggga agagattttt gaggaatcat atcgacaggt catgaaaatg agaccaaaag
     2161 atctctggaa gcgattaatg ataaaatttc gtggagaaga aggccttgac tatggaggcg
     2221 ttgccaggga atggttgtat ctcttgtcac atgaaatgtt gaatccatac tatggcctct
     2281 tccagtattc aagagatgat atttatacat tgcagatcaa tcctgattct gcagttaatc
     2341 cggaacattt atcctatttc cactttgttg gacgaataat gggaatggct gtgtttcatg
     2401 gacattatat tgatggtggt ttcacattgc ctttttataa gcaattgctt gggaagtcaa
     2461 ttaccttgga tgacatggag ttagtagatc cggatcttca caacagttta gtgtggatac
     2521 ttgagaatga tattacaggt gttttggacc ataccttctg tgttgaacat aatgcatatg
     2581 gtgaaattat tcagcatgaa cttaaaccaa atggcaaaag tatccctgtt aatgaagaaa
     2641 ataaaaaaga atatgtcagg ctctatgtga actggagatt tttacgaggc attgaggctc
     2701 aattcttggc tctgcagaaa ggatttaatg aagtaattcc acaacatctg ctgaagacat
     2761 ttgatgagaa ggagttagag ctcattattt gtggacttgg aaagatagat gttaatgact
     2821 ggaaggtaaa cacccggtta aaacactgta caccagacag caacattgtc aaatggttct
     2881 ggaaagctgt ggagtttttt gatgaagagc gacgagcaag attgcttcag tttgtgacag
     2941 gatcctctcg agtgcctctg cagggcttca aagcattgca aggtgctgca ggcccgagac
     3001 tctttaccat acaccagatt gatgcctgca ctaacaacct gccgaaagcc cacacttgct
     3061 tcaatcgaat agacattcca ccctatgaaa gctatgaaaa gctatatgaa aagctgctaa
     3121 cagccattga agaaacatgt ggatttgctg tggaatgaca atctagagcg gccgcatcgt
     3181 gactgactga cgatctgcct cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg
     3241 cagctcccgg agacggtcac agcttgtctg taagcggatg ccgggagcag acaagcccgt
     3301 cagggcgcgt cagcgggtgt tggcgggtgt cggggcgcag ccatgaccca gtcacgtagc
     3361 gatagcggag tgtataattc ttgaagacga aagggcctcg tgatacgcct atttttatag
     3421 gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg
     3481 cgcggaaccc ctatttgttt 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 acgttttcca
     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 cgatgcctgc 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 ttgatttaaa acttcatttt
     4501 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     4561 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     4621 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     4681 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     4741 agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag
     4801 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     4861 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     4921 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     4981 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     5041 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     5101 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     5161 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     5221 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     5281 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     5341 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg
     5401 tattttctcc ttacgcatct gtgcggtatt tcacaccgca taaattccga caccatcgaa
     5461 tggcgcaaaa cctttcgcgg tatggcatga tagcgcccgg aagagagtca attcagggtg
     5521 gtgaatgtga aaccagtaac gttatacgat gtcgcagagt atgccggtgt ctcttatcag
     5581 accgtttccc gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg ggaaaaagtg
     5641 gaagcggcga tggcggagct gaattacatt cccaaccgcg tggcacaaca actggcgggc
     5701 aaacagtcgt tgctgattgg cgttgccacc tccagtctgg ccctgcacgc gccgtcgcaa
     5761 attgtcgcgg cgattaaatc tcgcgccgat caactgggtg ccagcgtggt ggtgtcgatg
     5821 gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct cgcgcaacgc
     5881 gtcagtgggc tgatcattaa ctatccgctg gatgaccagg atgccattgc tgtggaagct
     5941 gcctgcacta atgttccggc gttatttctt gatgtctctg accagacacc catcaacagt
     6001 attattttct cccatgaaga cggtacgcga ctgggcgtgg agcatctggt cgcattgggt
     6061 caccagcaaa tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg tctgcgtctg
     6121 gctggctggc ataaatatct cactcgcaat caaattcagc cgatagcgga acgggaaggc
     6181 gactggagtg ccatgtccgg ttttcaacaa accatgcaaa tgctgaatga gggcatcgtt
     6241 cccactgcga tgctggttgc caacgatcag atggcgctgg gcgcaatgcg cgccattacc
     6301 gagtccgggc tgcgcgttgg tgcggatatc tcggtagtgg gatacgacga taccgaagac
     6361 agctcatgtt atatcccgcc gttaaccacc atcaaacagg attttcgcct gctggggcaa
     6421 accagcgtgg accgcttgct gcaactctct cagggccagg cggtgaaggg caatcagctg
     6481 ttgcccgtct cactggtgaa aagaaaaacc accctggcgc ccaatacgca aaccgcctct
     6541 ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc
     6601 gggcagtgag cgcaacgcaa ttaatgtaag ttagctcact cattaggcac cccaggcttt
     6661 acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac
     6721 aggaaacagc tatgaccatg attacggatt cactggccgt cgttttacaa cgtcgtgact
     6781 gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct ttcgccagct
     6841 ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc agcctgaatg
     6901 gcgaatggcg ctttgcctgg tttccggcac cagaagcggt gccggaaagc tggctggagt
     6961 gcgatcttcc tgaggccgat actgtcgtcg tcccctcaaa ctggcagatg cacggttacg
     7021 atgcgcccat ctacaccaac gtgacctatc ccattacggt caatccgccg tttgttccca
     7081 cggagaatcc gacgggttgt tactcgctca catttaatgt tgatgaaagc tggctacagg
     7141 aaggccagac gcgaattatt tttgatggcg ttggaattag cttatcgact gcacggtgca
     7201 ccaatgcttc tggcgtcagg cagccatcgg aagctgtggt atggctgtgc aggtcgtaaa
     7261 tcactgcata attcgtgtcg
//
