LOCUS       sequence-266384-        5962 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 5962)
  AUTHORS   Zhang Y, Feng XH, Derynck R
  TITLE     Smad3 and Smad4 cooperate with c-Jun/c-Fos to mediate 
            TGF-beta-induced transcription.
  JOURNAL   Nature. 1998 Aug 27. 394(6696):909-13.
  PUBMED    9732876
REFERENCE   2  (bases 1 to 5962)
  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. 
            1998 Aug 27. 394(6696):909-13."
COMMENT     Sequence Label: pRK5F Smad3
FEATURES             Location/Qualifiers
     source          1..5962
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        14..393
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        394..597
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     548..568
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     594..618
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        828..846
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     828..845
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     859..878
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             924..947
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     regulatory      2262..2271
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    2275..2409
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2312..2331)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2366..2385
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        2478..2835
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2478..2669
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2501..2521)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2686..2821
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2855..2872)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2855..2871)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2864..2886)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3084..3539
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3171..3190)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3381..3402
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3556..3575)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3675..3697
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3735..3753)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3821..3925
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3926..3994,3995..4786)
                     /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     complement(4144..4163)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4957..5545
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5446..5465
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5699..5716
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5833..5854
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5869..5874,5875..5892,5893..5899)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5907..5923
                     /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     5912..5934
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5931..5947
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5931..5947
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ttcgagctcg cccgacattg attattgact agttattaat agtaatcaat tacggggtca
       61 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
      121 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      181 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      241 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      301 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      361 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
      421 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
      481 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
      541 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctcgt
      601 ttagtgaacc gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga
      661 caccgggacc gatccagcct ccgcggccgg gaacggtgca ttggaacgcg gattccccgt
      721 gccaagagtg acgtaagtac cgcctataga gtctataggc ccaccccctt ggcttcgtta
      781 gaacgcggct acaattaata cataacctta tgtatcatac acatacgatt taggtgacac
      841 tatagaataa catccacttt gcctttctct ccacaggtgt ccactcccag gtccaactgc
      901 acctcggttc tatcgatacc atggactaca aggacgacga tgacaagggt cgacaatcgt
      961 ccatcctgcc tttcactccc ccgatcgtga agcgcctgct gggctggaag aagggcgagc
     1021 agaacgggca ggaggagaaa tggtgcgaga aggcggtcaa gagcctggtc aagaaactca
     1081 agaagacggg gcagctggac gagctggaga aggccatcac cacgcagaac gtcaacacca
     1141 agtgcatcac catccccagg tccctggatg gccggttgca ggtgtcccat cggaaggggc
     1201 tccctcatgt catctactgc cgcctgtggc gatggccaga cctgcacagc caccacgagc
     1261 tgcgggccat ggagctgtgt gagttcgcct tcaatatgaa gaaggacgag gtctgcgtga
     1321 atccctacca ctaccagaga gtagagacac cagttctacc tcctgtgttg gtgccacgcc
     1381 acacagagat cccggccgag ttccccccac tggacgacta cagccattcc atccccgaaa
     1441 acactaactt ccccgcaggc atcgagcccc agagcaatat tccagagacc ccaccccctg
     1501 gctacctgag tgaagatgga gaaaccagtg accaccagat gaaccacagc atggacgcag
     1561 gttctccaaa cctatccccg aatccgatgt ccccagcaca taataacttg gacctgcagc
     1621 cagttaccta ctgcgagccg gccttctggt gctccatctc ctactacgag ctgaaccagc
     1681 gcgtcgggga gacattccac gcctcgcagc catccatgac tgtggatggc ttcaccgacc
     1741 cctccaattc ggagcgcttc tgcctagggc tgctctccaa tgtcaacagg aatgcagcag
     1801 tggagctgac acggagacac atcggaagag gcgtgcggct ctactacatc ggaggggagg
     1861 tcttcgcaga gtgcctcagt gacagcgcta tttttgtcca gtctcccaac tgtaaccagc
     1921 gctatggctg gcacccggcc accgtctgca agatcccacc aggatgcaac ctgaagatct
     1981 tcaacaacca ggagttcgct gccctcctgg cccagtcggt caaccagggc tttgaggctg
     2041 tctaccagtt gacccgaatg tgcaccatcc gcatgagctt cgtcaaaggc tggggagcgg
     2101 agtacaggag acagactgtg accagtaccc cctgctggat tgagctgcac ctgaatgggc
     2161 ctttgcagtg gcttgacaag gtcctcaccc agatgggctc cccaagcatc cgctgttcca
     2221 gtgtgtctta ggatcctcta gagtcgacct gcagaagctt ggccgccatg gcccaacttg
     2281 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     2341 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     2401 gtctggatcg ggaattaatt cggcgcagca ccatggcctg aaataacctc tgaaagagga
     2461 acttggttag gtaccttctg aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg
     2521 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     2581 tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     2641 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact
     2701 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     2761 gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     2821 ctaggctttt gcaaaaagct gttaacagct tggcactggc cgtcgtttta caacgtcgtg
     2881 actgggaaaa ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca
     2941 gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctga
     3001 atggcgaatg gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
     3061 gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     3121 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     3181 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     3241 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgattt
     3301 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     3361 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     3421 ctcgggctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     3481 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt
     3541 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc
     3601 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     3661 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     3721 cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat
     3781 ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     3841 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     3901 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     3961 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     4021 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     4081 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     4141 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
     4201 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     4261 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     4321 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     4381 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     4441 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
     4501 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     4561 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     4621 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa
     4681 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     4741 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     4801 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     4861 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     4921 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     4981 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     5041 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     5101 tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     5161 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     5221 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     5281 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     5341 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     5401 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     5461 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     5521 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     5581 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     5641 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     5701 cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg
     5761 ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga
     5821 gcgcaacgca attaatgtga gttagctcac tcattaggca ccccaggctt tacactttat
     5881 gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag
     5941 ctatgacatg attacgaatt aa
//
