LOCUS       sequence-269493-        5240 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 5240)
  AUTHORS   Gao S, Alarcon C, Sapkota G, Rahman S, Chen PY, Goerner N, Macias 
            MJ, Erdjument-Bromage H, Tempst P, Massague J
  TITLE     Ubiquitin ligase Nedd4L targets activated Smad2/3 to limit TGF-beta 
            signaling.
  JOURNAL   Mol Cell. 2009 Nov 13. 36(3):457-68.
  PUBMED    19917253
REFERENCE   2  (bases 1 to 5240)
  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. 
            2009 Nov 13. 36(3):457-68."
COMMENT     Sequence Label: pCS2 Flag Smad3 S204A
FEATURES             Location/Qualifiers
     source          1..5240
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        469..487
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     469..486
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             540..563
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     polyA_signal    1933..2067
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1957..1976)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2011..2030
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2184..2204)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2184..2202)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2223..2239)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2223..2239)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2236..2258)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2247..2263
                     /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(2271..2277,2278..2295,2296..2301))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2316..2337
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2454..2471)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2625..3213)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2705..2724)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3384..4175,4176..4244))
                     /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     4007..4026
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4245..4349)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4375..4830)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4512..4533)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4724..4743
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 tatgggcggt cctattgacg tatatggcgc ctcccccatt gacgtcaatt acggtaaatg
       61 gcccgcctgg ctcaatgccc attgacgtca ataggaccac ccaccattga cgtcaatggg
      121 atggctcatt gcccattcat atccgttctc acgcccccta ttgacgtcaa tgacggtaaa
      181 tggcccactt ggcagtacat caatatctat taatagtaac ttggcaagta cattactatt
      241 ggaagtacgc cagggtacat tggcagtact cccattgacg tcaatggcgg taaatggccc
      301 gcgatggctg ccaagtacat ccccattgac gtcaatgggg aggggcaatg acgcaaatgg
      361 gcgttccatt gacgtaaatg ggcggtaggc gtgcctaatg ggaggtctat ataagcaatg
      421 ctcgtttagg gaaccgccat tctgcctggg gacgtcggag caagcttgat ttaggtgaca
      481 ctatagaata caagctactt gttctttttg caggatccca tcgattcgaa ttcaccatgg
      541 actacaagga cgacgatgac aagtcgtcca tcctgccttt cactcccccg atcgtgaagc
      601 gcctgctggg ctggaagaag ggcgagcaga acgggcagga ggagaaatgg tgcgagaagg
      661 cggtcaagag cctggtcaag aaactcaaga agacggggca gctggacgag ctggagaagg
      721 ccatcaccac gcagaacgtc aacaccaagt gcatcaccat ccccaggtcc ctggatggcc
      781 ggttgcaggt gtcccatcgg aaggggctcc ctcatgtcat ctactgccgc ctgtggcgat
      841 ggccagacct gcacagccac cacgagctgc gggccatgga gctgtgtgag ttcgccttca
      901 atatgaagaa ggacgaggtc tgcgtgaatc cctaccacta ccagagagta gagacaccag
      961 ttctacctcc tgtgttggtg ccacgccaca cagagatccc ggccgagttc cccccactgg
     1021 acgactacag ccattccatc cccgaaaaca ctaacttccc cgcaggcatc gagccccaga
     1081 gcaatattcc agagacccca ccccctggct acctgagtga agatggagaa accagtgacc
     1141 accagatgaa ccacagcatg gacgcaggtg ctccaaacct atccccgaat ccgatgtccc
     1201 cagcacataa taacttggac ctgcagccag ttacctactg cgagccggcc ttctggtgct
     1261 ccatctccta ctacgagctg aaccagcgcg tcggggagac attccacgcc tcgcagccat
     1321 ccatgactgt ggatggcttc accgacccct ccaattcgga gcgcttctgc ctagggctgc
     1381 tctccaatgt caacaggaat gcagcagtgg agctgacacg gagacacatc ggaagaggcg
     1441 tgcggctcta ctacatcgga ggggaggtct tcgcagagtg cctcagtgac agcgctattt
     1501 ttgtccagtc tcccaactgt aaccagcgct atggctggca cccggccacc gtctgcaaga
     1561 tcccaccagg atgcaacctg aagatcttca acaaccagga gttcgctgcc ctcctggccc
     1621 agtcggtcaa ccagggcttt gaggctgtct accagttgac ccgaatgtgc accatccgca
     1681 tgagcttcgt caaaggctgg ggagcggagt acaggagaca gactgtgacc agtaccccct
     1741 gctggattga gctgcacctg aatgggcctt tgcagtggct tgacaaggtc ctcacccaga
     1801 tgggctcccc aagcatccgc tgttccagtg tgtcttagag acatcgggct gcaggaattc
     1861 gtcgacaggc caatctggcc gcggccgcaa ggcctctcga gcctctagaa ctatagtgag
     1921 tcgtattacg tagatccaga catgataaga tacattgatg agtttggaca aaccacaact
     1981 agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta
     2041 accattataa gctgcaataa acaagttaac aacaacaatt gcattcattt tatgtttcag
     2101 gttcaggggg aggtgtggga ggttttttaa ttcgcggcgc gccgcggcgc caatgcattg
     2161 ggcccggtac ccagcttttg ttccctttag tgagggttaa ttgcgcgctt ggcgtaatca
     2221 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     2281 gccgggagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     2341 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     2401 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     2461 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     2521 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     2581 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     2641 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     2701 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     2761 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     2821 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     2881 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     2941 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     3001 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     3061 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     3121 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     3181 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     3241 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     3301 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     3361 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     3421 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     3481 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     3541 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     3601 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     3661 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     3721 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     3781 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     3841 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     3901 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     3961 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     4021 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     4081 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     4141 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     4201 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     4261 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     4321 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctaaattg
     4381 taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta
     4441 accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt
     4501 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca
     4561 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa
     4621 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat
     4681 ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag
     4741 gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg
     4801 ccgcgcttaa tgcgccgcta cagggcgcgt cccattcgcc attcaggctg cgcaactgtt
     4861 gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gtcgatcgac cgccaattca
     4921 atatggcgta tatggactca tgccaattca atatggtgga tctggacctg tgccaattca
     4981 atatggcgta tatggactcg tgccaattca atatggtgga tctggacccc agccaattca
     5041 atatggctat tggccaggtt caatactatg tattggccct atgccatata gtattccata
     5101 tatgggtttt cctattgacg tagatagccc ctcccaatgg gcggtcccat ataccatata
     5161 tggggcttcc taataccgcc catagccact cccccattga cgtcaatggt ctctatatat
     5221 ggtctttcct attgacgtca
//
