LOCUS       sequence-404933-        5187 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 5187)
  AUTHORS   Kretzschmar M, Doody J, Timokhina I, Massague J
  TITLE     A mechanism of repression of TGFbeta/ Smad signaling by oncogenic 
            Ras.
  JOURNAL   Genes Dev. 1999 Apr 1. 13(7):804-16.
  PUBMED    10197981
REFERENCE   2  (bases 1 to 5187)
  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: "Genes Dev. 
            1999 Apr 1. 13(7):804-16."
COMMENT     Sequence Label: CS2 Flag-Smad3 (3A)
FEATURES             Location/Qualifiers
     source          1..5187
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        453..471
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     453..470
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             524..547
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     polyA_signal    1864..1998
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1888..1907)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1942..1961
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2115..2135)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2115..2133)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2154..2170)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2154..2170)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2167..2189)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2178..2194
                     /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(2202..2208,2209..2226,2227..2232))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2247..2268
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2385..2402)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2556..3144)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2636..2655)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3315..4106,4107..4175))
                     /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     3938..3957
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4176..4280)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4306..4761)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4443..4464)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4655..4674
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gacgtatatg gcgcctcccc cattgacgtc aattacggta aatggcccgc ctggctcaat
       61 gcccattgac gtcaatagga ccacccacca ttgacgtcaa tgggatggct cattgcccat
      121 tcatatccgt tctcacgccc cctattgacg tcaatgacgg taaatggccc acttggcagt
      181 acatcaatat ctattaatag taacttggca agtacattac tattggaagt acgccagggt
      241 acattggcag tactcccatt gacgtcaatg gcggtaaatg gcccgcgatg gctgccaagt
      301 acatccccat tgacgtcaat ggggaggggc aatgacgcaa atgggcgttc cattgacgta
      361 aatgggcggt aggcgtgcct aatgggaggt ctatataagc aatgctcgtt tagggaaccg
      421 ccattctgcc tggggacgtc ggagcaagct tgatttaggt gacactatag aatacaagct
      481 acttgttctt tttgcaggat cccatcgatt cgaattcacc atggactaca aggacgacga
      541 tgacaagtcg tccatcctgc ctttcactcc cccgatcgtg aagcgcctgc tgggctggaa
      601 gaagggcgag cagaacgggc aggaggagaa atggtgcgag aaggcggtca agagcctggt
      661 caagaaactc aagaagacgg ggcagctgga cgagctggag aaggccatca ccacgcagaa
      721 cgtcaacacc aagtgcatca ccatccccag gtccctggat ggccggttgc aggtgtccca
      781 tcggaagggg ctccctcatg tcatctactg ccgcctgtgg cgatggccag acctgcacag
      841 ccaccacgag ctgcgggcca tggagctgtg tgagttcgcc ttcaatatga agaaggacga
      901 ggtctgcgtg aatccctacc actaccagag agtagagaca ccagttctac ctcctgtgtt
      961 ggtgccacgc cacacagaga tcccggccga gttcccccca ctggacgact acagccattc
     1021 catccccgaa aacactaact tccccgcagg catcgagccc cagagcaata ttccagagac
     1081 cccaccccct ggctacctga gtgaagatgg agaaaccagt gaccaccaga tgaaccacag
     1141 catggacgca ggttctccaa acctatcccc gaatccgatg tccccagcac ataataactt
     1201 ggacctgcag ccagttacct actgcgagcc ggccttctgg tgctccatct cctactacga
     1261 gctgaaccag cgcgtcgggg agacattcca cgcctcgcag ccatccatga ctgtggatgg
     1321 cttcaccgac ccctccaatt cggagcgctt ctgcctaggg ctgctctcca atgtcaacag
     1381 gaatgcagca gtggagctga cacggagaca catcggaaga ggcgtgcggc tctactacat
     1441 cggaggggag gtcttcgcag agtgcctcag tgacagcgct atttttgtcc agtctcccaa
     1501 ctgtaaccag cgctatggct ggcacccggc caccgtctgc aagatcccac caggatgcaa
     1561 cctgaagatc ttcaacaacc aggagttcgc tgccctcctg gcccagtcgg tcaaccaggg
     1621 ctttgaggct gtctaccagt tgacccgaat gtgcaccatc cgcatgagct tcgtcaaagg
     1681 ctggggagcg gagtacagga gacagactgt gaccagtacc ccctgctgga ttgagctgca
     1741 cctgaatggg cctttgcagt ggcttgacaa ggtcctcacc cagatgggct ccccaagcat
     1801 ccgctgtgcc gctgtggctt agggatctcg agcctctaga actatagtga gtcgtattac
     1861 gtagatccag acatgataag atacattgat gagtttggac aaaccacaac tagaatgcag
     1921 tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt aaccattata
     1981 agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca ggttcagggg
     2041 gaggtgtggg aggtttttta attcgcggcg cgccgcggcg ccaatgcatt gggcccggta
     2101 cccagctttt gttcccttta gtgagggtta attgcgcgct tggcgtaatc atggtcatag
     2161 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccgggagc
     2221 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     2281 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     2341 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     2401 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     2461 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     2521 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     2581 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     2641 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     2701 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     2761 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     2821 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     2881 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     2941 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     3001 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     3061 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     3121 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     3181 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     3241 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     3301 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     3361 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     3421 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     3481 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     3541 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     3601 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     3661 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     3721 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     3781 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     3841 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     3901 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     3961 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     4021 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     4081 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     4141 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     4201 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     4261 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctaaatt gtaagcgtta
     4321 atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg
     4381 ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg
     4441 ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa
     4501 aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg
     4561 ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt
     4621 gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg
     4681 ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta
     4741 atgcgccgct acagggcgcg tcccattcgc cattcaggct gcgcaactgt tgggaagggc
     4801 gatcggtgcg ggcctcttcg ctattacgcc agtcgatcga ccgccaattc aatatggcgt
     4861 atatggactc atgccaattc aatatggtgg atctggacct gtgccaattc aatatggcgt
     4921 atatggactc gtgccaattc aatatggtgg atctggaccc cagccaattc aatatggcta
     4981 ttggccaggt tcaatactat gtattggccc tatgccatat agtattccat atatgggttt
     5041 tcctattgac gtagatagcc cctcccaatg ggcggtccca tataccatat atggggcttc
     5101 ctaataccgc ccatagccac tcccccattg acgtcaatgg tctctatata tggtctttcc
     5161 tattgacgtc atatgggcgg tcctatt
//
