LOCUS       sequence-315121-        7009 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  encodes transcriptional reporter of SMAD2/3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7009)
  AUTHORS   Taylor B, Shah A, Bielczyk-Maczynska E
  TITLE     TGF-beta is insufficient to induce adipocyte state loss without 
            concurrent PPARgamma downregulation.
  JOURNAL   Sci Rep. 2020 Aug 21;10(1):14084. doi: 10.1038/s41598-020-71100-z.
  PUBMED    32826933
REFERENCE   2  (bases 1 to 7009)
  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: "Sci Rep."; 
            date: "2020-08-21"; volume: "10(1):14084. doi"; pages: " 
            10.1038/s41598-020-71100-z"
COMMENT     Sequence Label: PB_SBE4:mScarlet-I-NLS
FEATURES             Location/Qualifiers
     source          1..7009
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..63
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     misc_feature    479..570
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     519..538
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     promoter        679..875
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      726..861
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     regulatory      905..914
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             911..1606
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet-I
                     /note="matures faster than mScarlet"
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFIKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(1044..1062)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1312..1331
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1610..1630
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     promoter        1645..2144
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(1680..1703)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2077..2096
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             2171..2569
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     primer_bind     complement(2644..2664)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    2706..2827
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2743..2762)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2797..2816
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   3590..3624
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(4115..4131)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4115..4131)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4128..4150)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4139..4155
                     /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(4163..4169,4170..4187,4188..4193))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4208..4229
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4346..4363)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4517..5105)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4597..4616)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5276..6067,6068..6136))
                     /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     5899..5918
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6137..6241)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6267..6722)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6404..6425)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6616..6635
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6849..6871
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6863..6880
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6864..6880
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 ccctagaaag ataatcatat tgtgacgtac gttaaagata atcatgtgta aaattgacgc
       61 atgtgtttta tcggtctgta tatcgaggtt tatttattaa tttgaataga tattaagttt
      121 tattatattt acacttacat actaataata aattcaacaa acaatttatt tatgtttatt
      181 tatttattaa aaaaaacaaa aactcaaaat ttcttctata aagtaacaaa acttttatga
      241 gggacagccc ccccccaaag cccccaggga tgtaattacg tccctccccc gctagggggc
      301 agcagcgagc cgcccggggc tccgctccgg tccggcgctc cccccgcatc cccgagccgg
      361 cagcgtgcgg ggacagcccg ggcacgggga aggtggcacg ggatcgcttt cctctgaacg
      421 cttctcgctg ctctttgagc ctgcagacac ctggggggat acggggaaaa ggcctccaaa
      481 catacgctct ccatcaaaac aaaacgaaac aaaacaaact agcaaaatag gctgtcccca
      541 gtgcaagtgc aggtgccaga acatttctct atcgataggt actaagtcta gacggcagtc
      601 tagacgtact aagtctagac ggcagtctag acgtaccgag ctcttacgcg tgctagcccg
      661 ggctcgagat ctgcgatctg catctcaatt agtcagcaac catagtcccg cccctaactc
      721 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat cgctgactaa
      781 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
      841 gaggaggctt ttttggaggc ctaggctttt gcaaaaagct tggcattccg gtactgttgg
      901 taaagccacc atggtgagca agggcgaggc agtgatcaag gagttcatgc ggttcaaggt
      961 gcacatggag ggctccatga acggccacga gttcgagatc gagggcgagg gcgagggccg
     1021 cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggtggcc ccctgccctt
     1081 ctcctgggac atcctgtccc ctcagttcat gtacggctcc agggccttca tcaagcaccc
     1141 cgccgacatc cccgactact ataagcagtc cttccccgag ggcttcaagt gggagcgcgt
     1201 gatgaacttc gaggacggcg gcgccgtgac cgtgacccag gacacctccc tggaggacgg
     1261 caccctgatc tacaaggtga agctccgcgg caccaacttc cctcctgacg gccccgtaat
     1321 gcagaagaag acaatgggct gggaagcgtc caccgagcgg ttgtaccccg aggacggcgt
     1381 gctgaagggc gacattaaga tggccctgcg cctgaaggac ggcggccgct acctggcgga
     1441 cttcaagacc acctacaagg ccaagaagcc cgtgcagatg cccggcgcct acaacgtcga
     1501 ccgcaagttg gacatcacct cccacaacga ggactacacc gtggtggaac agtacgaacg
     1561 ctccgagggc cgccactcca ccggcggcat ggacgagctg tacaagcctc caaaaaagaa
     1621 gagaaaggta tagagaattc taccgggtag gggaggcgct tttcccaagg cagtctggag
     1681 catgcgcttt agcagccccg ctgggcactt ggcgctacac aagtggcctc tggcctcgca
     1741 cacattccac atccaccggt aggcgccaac cggctccgtt ctttggtggc cccttcgcgc
     1801 caccttctac tcctccccta gtcaggaagt tcccccccgc cccgcagctc gcgtcgtgca
     1861 ggacgtgaca aatggaagta gcacgtctca ctagtctcgt gcagatggac agcaccgctg
     1921 agcaatggaa gcgggtaggc ctttggggca gcggccaata gcagctttgc tccttcgctt
     1981 tctgggctca gaggctggga aggggtgggt ccgggggcgg gctcaggggc gggctcaggg
     2041 gcggggcggg cgcccgaagg tcctccggag gcccggcatt ctgcacgctt caaaagcgca
     2101 cgtctgccgc gctgttctcc tcttcctcat ctccgggcct ttcgacctgc agcccaagct
     2161 tcctaggacc atggccaagc ctttgtctca agaagaatcc accctcattg aaagagcaac
     2221 ggctacaatc aacagcatcc ccatctctga agactacagc gtcgccagcg cagctctctc
     2281 tagcgacggc cgcatcttca ctggtgtcaa tgtatatcat tttactgggg gaccttgtgc
     2341 agaactcgtg gtgctgggca ctgctgctgc tgcggcagct ggcaacctga cttgtatcgt
     2401 cgcgatcgga aatgagaaca ggggcatctt gagcccctgc ggacggtgcc gacaggtgct
     2461 tctcgatctg catcctggga tcaaagccat agtgaaggac agtgatggac agccgacggc
     2521 agttgggatt cgtgaattgc tgccctctgg ttatgtgtgg gagggctaac ccgggcgggg
     2581 cgcgtctggc aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa
     2641 ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgcgtt
     2701 aactaaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     2761 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     2821 tatcttatca tgtctggaat tgactcaaat gatgtcaatt agtctatcag aagctcatct
     2881 ggtctccctt ccgggggaca agacatccct gtttaatatt taaacagcag tgttcccaaa
     2941 ctgggttctt atatcccttg ctctggtcaa ccaggttgca gggtttcctg tcctcacagg
     3001 aacgaagtcc ctaaagaaac agtggcagcc aggtttagcc ccggaattga ctggattcct
     3061 tttttagggc ccattggtat ggctttttcc ccgtatcccc ccaggtgtct gcaggctcaa
     3121 agagcagcga gaagcgttca gaggaaagcg atcccgtgcc accttccccg tgcccgggct
     3181 gtccccgcac gctgccggct cggggatgcg gggggagcgc cggaccggag cggagccccg
     3241 ggcggctcgc tgctgccccc tagcggggga gggacgtaat tacatccctg ggggctttgg
     3301 gggggggctg tccctgatat ctataacaag aaaatatata tataataagt tatcacgtaa
     3361 gtagaacatg aaataacaat ataattatcg tatgagttaa atcttaaaag tcacgtaaaa
     3421 gataatcatg cgtcattttg actcacgcgg tcgttatagt tcaaaatcag tgacacttac
     3481 cgcattgaca agcacgcctc acgggagctc caagcggcga ctgagatgtc ctaaatgcac
     3541 agcgacggat tcgcgctatt tagaaagaga gagcaatatt tcaagaatgc atgcgtcaat
     3601 tttacgcaga ctatctttct agggttaatc tagctgcatc aggatcatat cgtcgggtct
     3661 tttttccggc tcagtcatcg cccaagctgg cgctatctgg gcatcgggga ggaagaagcc
     3721 cgtgcctttt cccgcgaggt tgaagcggca tggaaagagt ttgccgagga tgactgctgc
     3781 tgcattgacg ttgagcgaaa acgcacgttt accatgatga ttcgggaagg tgtggccatg
     3841 cacgccttta acggtgaact gttcgttcag gccacctggg ataccagttc gtcgcggctt
     3901 ttccggacac agttccggat ggtcagcccg aagcgcatca gcaacccgaa caataccggc
     3961 gacagccgga actgccgtgc cggtgtgcag attaatgaca gcggtgcggc gctgggatat
     4021 tacgtcagcg aggacgggta tcctggctgg atgccgcaga aatggacatg gataccccgt
     4081 gagttacccg gcgggcgcgc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
     4141 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg
     4201 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
     4261 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     4321 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     4381 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     4441 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     4501 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     4561 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     4621 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     4681 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     4741 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     4801 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     4861 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     4921 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     4981 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     5041 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     5101 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     5161 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     5221 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     5281 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     5341 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     5401 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     5461 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     5521 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     5581 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     5641 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     5701 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     5761 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     5821 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     5881 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     5941 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     6001 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     6061 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     6121 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     6181 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     6241 gcacatttcc ccgaaaagtg ccacctaaat tgtaagcgtt aatattttgt taaaattcgc
     6301 gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc
     6361 ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag
     6421 tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga
     6481 tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc
     6541 actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa
     6601 cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt
     6661 agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc
     6721 gtcccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc
     6781 gctattacgc cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc
     6841 agggttttcc cagtcacgac gttgtaaaac gacggccagt gagcgcgcct cgttcattca
     6901 cgtttttgaa cccgtggagg acgggcagac tcgcggtgca aatgtgtttt acagcgtgat
     6961 ggagcagatg aagatgctcg acacgctgca gaacacgcag ctagattaa
//
