LOCUS       sequence-454192-        6847 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 6847)
  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 6847)
  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: pCMV5B-Flag-Smurf1 wt
FEATURES             Location/Qualifiers
     source          1..6847
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     94..116
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     108..125
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     109..125
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        286..665
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        666..869
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     820..840
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             921..944
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     polyA_signal    3156..3778
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3355..3374)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(3802..3822)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3807..4136
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3987..4122
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4174..4193)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4175..4193)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4207..4223)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4207..4223)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4220..4242)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4231..4247
                     /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(4255..4261,4262..4279,4280..4285))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4300..4321
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4438..4455)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4609..5197)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4689..4708)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5368..6159,6160..6228))
                     /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     5991..6010
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6229..6333)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6360..6815
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6447..6466)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6657..6678
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 aagggcgatc ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg
       61 caaggcgatt aagttgggta acgccagggt tttcccagtc acgacgttgt aaaacgacgg
      121 ccagtgccaa gctgatctat acattgaatc aatattggca attagccata ttagtcattg
      181 gttatatagc ataaatcaat attggctatt ggccattgca tacgttgtat ctatatcata
      241 atatgtacat ttatattggc tcatgtccaa tatgaccgcc atgttgacat tgattattga
      301 ctagttatta atagtaatca attacggggt cattagttca tagcccatat atggagttcc
      361 gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat
      421 tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc
      481 aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc
      541 caagtccgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt
      601 acatgacctt acgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta
      661 ccatggtgat gcggttttgg cagtacacca atgggcgtgg atagcggttt gactcacggg
      721 gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac
      781 gggactttcc aaaatgtcgt aataaccccg ccccgttgac gcaaatgggc ggtaggcgtg
      841 tacggtggga ggtctatata agcagagctc gtttagtgaa ccgtcagaat tgatctggta
      901 ccacgcgtat cgataccatg gactacaagg acgacgatga caagggtcga caaagcaacg
      961 tggtgacacg gagaggaggc tccagcatca agatccgtct gacagtgtta tgtgccaaga
     1021 accttgcaaa gaaagacttc ttcaggctcc ctgacccttt tgcaaagatt gtcgtggatg
     1081 ggtctgggca gtgccactca accgacactg tgaaaaacac attggaccca aagtggaacc
     1141 agcactatga tctatatgtt gggaaaacgg attcgataac cattagcgtg tggaaccata
     1201 agaaaattca caagaaacag ggagctggct tcctgggctg tgtgcggctg ctctccaatg
     1261 ccatcagcag attaaaagat accggatacc agcgtttgga tctatgcaaa ctaaacccct
     1321 cagatactga tgcagttcgt ggccagatag tggtcagttt acagacacga gacagaatag
     1381 gaaccggcgg ctcggtggtg gactgcagag gactgttaga aaatgaagga acggtgtatg
     1441 aagactccgg gcctgggagg ccgctcagct gcttcatgga ggaaccagcc ccttacacag
     1501 atagcaccgg tgctgctgct ggaggaggga attgcaggtt cgtggagtcc ccaagtcaag
     1561 atcaaagact tcaggcacag cggcttcgaa accctgatgt gcgaggttca ctacagacgc
     1621 cccagaaccg accacacggc caccagtccc cggaactgcc cgaaggctac gaacaaagaa
     1681 caacagtcca gggccaagtt tactttttgc atacacagac tggagttagc acgtggcacg
     1741 accccaggat accaagagac cttaacagtg tgaactgtga tgaacttgga ccactgccgc
     1801 caggctggga agtcagaagt acagtttctg ggaggatata ttttgtagat cataataacc
     1861 gaacaaccca gtttacagac ccaaggttac accacatcat gaatcaccag tgccaactca
     1921 aggagcccag ccagccgctg ccactgccca gtgagggctc tctggaggac gaggagcttc
     1981 ctgcccagag atacgaaaga gatctagtcc agaagctgaa agtcctcaga cacgaactgt
     2041 cgcttcagca gccccaagct ggtcattgcc gcatcgaagt gtccagagaa gaaatctttg
     2101 aggagtctta ccgccagata atgaagatgc gaccgaaaga cttgaaaaaa cggctgatgg
     2161 tgaaattccg tggggaagaa ggtttggatt acggtggtgt ggccagggag tggctttact
     2221 tgctgtgcca tgaaatgctg aatccttatt acgggctctt ccagtattct acggacaata
     2281 tttacatgtt gcaaataaat ccggattctt caatcaaccc cgaccacttg tcttatttcc
     2341 actttgtggg gcggatcatg gggctggctg tgttccatgg acactacatc aacgggggct
     2401 tcacagtgcc cttctacaag cagctgctgg ggaagcccat ccagctctca gatctggaat
     2461 ctgtggaccc agagctgcat aagagcttgg tgtggatcct agagaacgac atcacgcctg
     2521 tactggacca caccttctgc gtggaacaca acgccttcgg gcggatcctg cagcatgaac
     2581 tgaaacccaa tggcagaaat gtgccagtca cagaggagaa taagaaagaa tacgtccggt
     2641 tgtatgtaaa ctggaggttt atgagaggaa tcgaagccca gttcttagct ctgcagaagg
     2701 ggttcaatga gctcatccct caacatctgc tgaagccttt tgaccagaag gaactggagc
     2761 tgatcatagg cggcctggat aaaatagact tgaacgactg gaagtcgaac acgcggctga
     2821 agcactgtgt ggccgacagc aacatcgtgc ggtggttctg gcaagcggtg gagacgttcg
     2881 atgaagaaag gagggccagg ctcctgcagt ttgtgactgg gtccacgcga gtcccgctcc
     2941 aaggcttcaa ggctttgcaa ggttctacag gcgcggcagg gccccggctg ttcaccatcc
     3001 acctgataga cgcgaacaca gacaaccttc cgaaggccca tacctgcttt aaccggatcg
     3061 acattccacc atatgagtcc tatgagaagc tctacgagaa gctgctgaca gccgtggagg
     3121 agacctgcgg gtttgctgtg gagtaatcta gacccgggtg gcatccctgt gacccctccc
     3181 cagtgcctct cctggccctg gaagttgcca ctccagtgcc caccagcctt gtcctaataa
     3241 aattaagttg catcattttg tctgactagg tgtccttcta taatattatg gggtggaggg
     3301 gggtggtatg gagcaagggg caagttggga agacaacctg tagggcctgc ggggtctatt
     3361 gggaaccaag ctggagtgca gtggcacaat cttggctcac tgcaatctcc gcctcctggg
     3421 ttcaagcgat tctcctgcct cagcctcccg agttgttggg attccaggca tgcatgacca
     3481 ggctcagcta atttttgttt ttttggtaga gacggggttt caccatattg gccaggctgg
     3541 tctccaactc ctaatctcag gtgatctacc caccttggcc tcccaaattg ctgggattac
     3601 aggcgtgaac cactgctccc ttccctgtcc ttctgatttt aaaataacta taccagcagg
     3661 aggacgtcca gacacagcat aggctacctg gccatgccca accggtggga catttgagtt
     3721 gcttgcttgg cactgtcctc tcatgcgttg ggtccactca gtagatgcct gttgaattgg
     3781 gtacgcggcc agcttggctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
     3841 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
     3901 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     3961 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     4021 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct
     4081 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc
     4141 tcctcgagga actgaaaaac cagaaagtta attccctata gtgagtcgta ttaaattcgt
     4201 aatcatgtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     4261 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     4321 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     4381 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     4441 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     4501 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     4561 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     4621 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     4681 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     4741 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     4801 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     4861 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     4921 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     4981 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     5041 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     5101 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     5161 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     5221 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     5281 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     5341 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     5401 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     5461 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     5521 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     5581 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     5641 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     5701 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     5761 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     5821 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     5881 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     5941 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     6001 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     6061 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     6121 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     6181 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     6241 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     6301 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     6361 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     6421 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     6481 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     6541 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     6601 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     6661 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     6721 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     6781 cgcgaatttt aacaaaatat taacgcttac aatttccatt cgccattcag gctgcgcaac
     6841 tgttggg
//
