LOCUS       sequence-313743-        6039 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 6039)
  AUTHORS   Lo RS, Massague J
  TITLE     Ubiquitin-dependent degradation of TGF-beta-activated smad2.
  JOURNAL   Nat Cell Biol. 1999 Dec . 1(8):472-8.
  PUBMED    10587642
REFERENCE   2  (bases 1 to 6039)
  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: "Nat Cell 
            Biol. 1999 Dec . 1(8):472-8."
COMMENT     Sequence Label: pCMV5 Flag-NLS-Smad2
FEATURES             Location/Qualifiers
     source          1..6039
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             619..642
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             649..669
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    2084..2706
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2283..2302)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(2730..2750)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2895..3029
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3081..3100)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3082..3100)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3114..3130)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3114..3130)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3127..3149)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3138..3154
                     /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(3162..3168,3169..3186,3187..3192))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3207..3228
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3345..3362)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3516..4104)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3596..3615)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4275..5066,5067..5135))
                     /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     4898..4917
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5136..5240)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5267..5722
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5354..5373)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5564..5585
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5848..5870
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5862..5879
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5863..5879
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt ccgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttacggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta caccaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaataa ccccgccccg ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc
      601 agaattgcag tgaccatgga ctacaaggac gacgatgaca aggaattccc caagaagaag
      661 agaaaggtgg agatgtcgtc catcttgcca ttcacgccgc cagttgtgaa gagactgctg
      721 ggatggaaga agtcagctgg tgggtctgga ggagcaggcg gaggagagca gaatgggcag
      781 gaagaaaagt ggtgtgagaa agcagtgaaa agttcggtga agaagctaaa gaaaacagga
      841 cgattagatg agcttgagaa agccatcacc actcaaaact gtaatactaa atgtgttacc
      901 ataccaagca cttgctctga aatttgggga ctgagtacac caaatacgat agatcagtgg
      961 gatacaacag gcctttacag cttctctgaa caaaccaggt ctcttgatgg tcgtctccag
     1021 gtatcccatc gaaaaggatt gccacatgtt atatattgcc gattatggcg ctggcctgat
     1081 cttcacagtc atcatgaact caaggcaatt gaaaactgcg aatatgcttt taatcttaaa
     1141 aaggatgaag tatgtgtaaa cccttaccac tatcagagag ttgagacacc agttttgcct
     1201 ccagtattag tgccccgaca caccgagatc ctaacagaac ttccgcctct ggatgactat
     1261 actcactcca ttccagaaaa cactaacttc ccagcaggaa ttgagccaca gagtaattat
     1321 attccagaaa cgccacctcc tggatatatc agtgaagatg gagaaacaag tgaccaacag
     1381 ttgaatcaaa gtatggacac aggctctcca gcagaactat ctcctactac tctttcccct
     1441 gttaatcata gcttggattt acagccagtt acttactcag aacctgcatt ttggtgttca
     1501 atagcatatt atgaattaaa tcagagggtt ggagaaacct tccatgcatc acagccctca
     1561 ctcactgtag atggctttac agacccatca aattcagaga ggttctgctt aggtttactc
     1621 tccaatgtta accgaaatgc cacggtagaa atgacaagaa ggcatatagg aagaggagtg
     1681 cgcttatact acataggtgg ggaagttttt gctgagtgcc taagtgatag tgcaatcttt
     1741 gtgcagagcc ccaattgtaa tcagagatat ggctggcacc ctgcaacagt gtgtaaaatt
     1801 ccaccaggct gtaatctgaa gatcttcaac aaccaggaat ttgctgctct tctggctcag
     1861 tctgttaatc agggttttga agccgtctat cagctaacta gaatgtgcac cataagaatg
     1921 agttttgtga aagggtgggg agcagaatac cgaaggcaga cggtaacaag tactccttgc
     1981 tggattgaac ttcatctgaa tggacctcta cagtggttgg acaaagtatt aactcagatg
     2041 ggatctcctt cagtgcgttg ctcaagcatg tcataaggat cccgggtggc atccctgtga
     2101 cccctcccca gtgcctctcc tggccctgga agttgccact ccagtgccca ccagccttgt
     2161 cctaataaaa ttaagttgca tcattttgtc tgactaggtg tccttctata atattatggg
     2221 gtggaggggg gtggtatgga gcaaggggca agttgggaag acaacctgta gggcctgcgg
     2281 ggtctattgg gaaccaagct ggagtgcagt ggcacaatct tggctcactg caatctccgc
     2341 ctcctgggtt caagcgattc tcctgcctca gcctcccgag ttgttgggat tccaggcatg
     2401 catgaccagg ctcagctaat ttttgttttt ttggtagaga cggggtttca ccatattggc
     2461 caggctggtc tccaactcct aatctcaggt gatctaccca ccttggcctc ccaaattgct
     2521 gggattacag gcgtgaacca ctgctccctt ccctgtcctt ctgattttaa aataactata
     2581 ccagcaggag gacgtccaga cacagcatag gctacctggc catgcccaac cggtgggaca
     2641 tttgagttgc ttgcttggca ctgtcctctc atgcgttggg tccactcagt agatgcctgt
     2701 tgaattgggt acgcggccag cttggctgtg gaatgtgtgt cagttagggt gtggaaagtc
     2761 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag
     2821 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     2881 gtcagcaacc atagtcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg
     2941 ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca
     3001 gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc tcgaggaact
     3061 gaaaaaccag aaagttaatt ccctatagtg agtcgtatta aattcgtaat catgtcatag
     3121 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     3181 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     3241 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     3301 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     3361 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     3421 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     3481 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     3541 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     3601 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     3661 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     3721 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     3781 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     3841 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     3901 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     3961 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     4021 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     4081 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     4141 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     4201 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     4261 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     4321 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     4381 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     4441 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     4501 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     4561 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     4621 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     4681 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     4741 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     4801 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     4861 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     4921 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     4981 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     5041 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     5101 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     5161 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     5221 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgc gccctgtagc
     5281 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     5341 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     5401 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     5461 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     5521 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     5581 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     5641 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac
     5701 aaaatattaa cgcttacaat ttccattcgc cattcaggct gcgcaactgt tgggaagggc
     5761 gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc
     5821 gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg
     5881 ccaagctgat ctatacattg aatcaatatt ggcaattagc catattagtc attggttata
     5941 tagcataaat caatattggc tattggccat tgcatacgtt gtatctatat cataatatgt
     6001 acatttatat tggctcatgt ccaatatgac cgccatgtt
//
