LOCUS       sequence-299917-        7001 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 7001)
  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 7001)
  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..7001
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(34..53)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(713..1504,1505..1573))
                     /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     1336..1355
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1574..1678)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(1704..2159)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1841..1862)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2053..2072
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2286..2308
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2300..2317
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2301..2317
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   2447..2509
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     misc_feature    2925..3016
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     2965..2984
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     promoter        3125..3321
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3172..3307
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     regulatory      3351..3360
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3357..4052
                     /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(3490..3508)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3758..3777
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             4056..4076
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     promoter        4091..4590
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(4126..4149)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     4523..4542
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             4617..5015
                     /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(5090..5110)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    5145..5266
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5182..5201)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5236..5255
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   6028..6062
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(6553..6569)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6553..6569)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6566..6588)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6577..6593
                     /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(6601..6607,6608..6625,6626..6631))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6646..6667
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6784..6801)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(6955..7001,1..542))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
       61 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
      121 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
      181 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
      241 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
      301 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
      361 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc
      421 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
      481 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
      541 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
      601 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
      661 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
      721 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
      781 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
      841 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
      901 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
      961 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     1021 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     1081 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     1141 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     1201 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     1261 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     1321 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     1381 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     1441 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     1501 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     1561 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     1621 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     1681 catttccccg aaaagtgcca cctaaattgt aagcgttaat attttgttaa aattcgcgtt
     1741 aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta
     1801 taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc
     1861 actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg
     1921 cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact
     1981 aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt
     2041 ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc
     2101 ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc
     2161 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
     2221 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
     2281 gttttcccag tcacgacgtt gtaaaacgac ggccagtgag cgcgcctcgt tcattcacgt
     2341 ttttgaaccc gtggaggacg ggcagactcg cggtgcaaat gtgttttaca gcgtgatgga
     2401 gcagatgaag atgctcgaca cgctgcagaa cacgcagcta gattaaccct agaaagataa
     2461 tcatattgtg acgtacgtta aagataatca tgcgtaaaat tgacgcatgt gttttatcgg
     2521 tctgtatatc gaggtttatt tattaatttg aatagatatt aagttttatt atatttacac
     2581 ttacatacta ataataaatt caacaaacaa tttatttatg tttatttatt tattaaaaaa
     2641 aacaaaaact caaaatttct tctataaagt aacaaaactt ttatgaggga cagccccccc
     2701 ccaaagcccc cagggatgta attacgtccc tcccccgcta gggggcagca gcgagccgcc
     2761 cggggctccg ctccggtccg gcgctccccc cgcatccccg agccggcagc gtgcggggac
     2821 agcccgggca cggggaaggt ggcacgggat cgctttcctc tgaacgcttc tcgctgctct
     2881 ttgagcctgc agacacctgg ggggatacgg ggaaaaggcc tccaaacata cgctctccat
     2941 caaaacaaaa cgaaacaaaa caaactagca aaataggctg tccccagtgc aagtgcaggt
     3001 gccagaacat ttctctatcg ataggtacta agtctagacg gcagtctaga cgtactaagt
     3061 ctagacggca gtctagacgt accgagctct tacgcgtgct agcccgggct cgagatctgc
     3121 gatctgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc
     3181 ctaactccgc ccagttccgc ccattctccg ccccatcgct gactaatttt ttttatttat
     3241 gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt
     3301 ggaggcctag gcttttgcaa aaagcttggc attccggtac tgttggtaaa gccaccatgg
     3361 tgagcaaggg cgaggcagtg atcaaggagt tcatgcggtt caaggtgcac atggagggct
     3421 ccatgaacgg ccacgagttc gagatcgagg gcgagggcga gggccgcccc tacgagggca
     3481 cccagaccgc caagctgaag gtgaccaagg gtggccccct gcccttctcc tgggacatcc
     3541 tgtcccctca gttcatgtac ggctccaggg ccttcatcaa gcaccccgcc gacatccccg
     3601 actactataa gcagtccttc cccgagggct tcaagtggga gcgcgtgatg aacttcgagg
     3661 acggcggcgc cgtgaccgtg acccaggaca cctccctgga ggacggcacc ctgatctaca
     3721 aggtgaagct ccgcggcacc aacttccctc ctgacggccc cgtaatgcag aagaagacaa
     3781 tgggctggga agcgtccacc gagcggttgt accccgagga cggcgtgctg aagggcgaca
     3841 ttaagatggc cctgcgcctg aaggacggcg gccgctacct ggcggacttc aagaccacct
     3901 acaaggccaa gaagcccgtg cagatgcccg gcgcctacaa cgtcgaccgc aagttggaca
     3961 tcacctccca caacgaggac tacaccgtgg tggaacagta cgaacgctcc gagggccgcc
     4021 actccaccgg cggcatggac gagctgtaca agcctccaaa aaagaagaga aaggtataga
     4081 gaattctacc gggtagggga ggcgcttttc ccaaggcagt ctggagcatg cgctttagca
     4141 gccccgctgg gcacttggcg ctacacaagt ggcctctggc ctcgcacaca ttccacatcc
     4201 accggtaggc gccaaccggc tccgttcttt ggtggcccct tcgcgccacc ttctactcct
     4261 cccctagtca ggaagttccc ccccgccccg cagctcgcgt cgtgcaggac gtgacaaatg
     4321 gaagtagcac gtctcactag tctcgtgcag atggacagca ccgctgagca atggaagcgg
     4381 gtaggccttt ggggcagcgg ccaatagcag ctttgctcct tcgctttctg ggctcagagg
     4441 ctgggaaggg gtgggtccgg gggcgggctc aggggcgggc tcaggggcgg ggcgggcgcc
     4501 cgaaggtcct ccggaggccc ggcattctgc acgcttcaaa agcgcacgtc tgccgcgctg
     4561 ttctcctctt cctcatctcc gggcctttcg acctgcagcc caagcttcct aggaccatgg
     4621 ccaagccttt gtctcaagaa gaatccaccc tcattgaaag agcaacggct acaatcaaca
     4681 gcatccccat ctctgaagac tacagcgtcg ccagcgcagc tctctctagc gacggccgca
     4741 tcttcactgg tgtcaatgta tatcatttta ctgggggacc ttgtgcagaa ctcgtggtgc
     4801 tgggcactgc tgctgctgcg gcagctggca acctgacttg tatcgtcgcg atcggaaatg
     4861 agaacagggg catcttgagc ccctgcggac ggtgccgaca ggtgcttctc gatctgcatc
     4921 ctgggatcaa agccatagtg aaggacagtg atggacagcc gacggcagtt gggattcgtg
     4981 aattgctgcc ctctggttat gtgtgggagg gctaacccgg gcggggcgcg tctggcaatc
     5041 aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat gttgctcctt
     5101 ttacgctatg tggatacgct gctttaatgc ctttgtatca tgttaacttg tttattgcag
     5161 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     5221 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctggaatt
     5281 gactcaaatg atgtcaatta gtctatcaga agctatctgg tctcccttcc gggggacaag
     5341 acatccctgt ttaatattta aacagcagtg ttcccaaact gggttcttat atcccttgct
     5401 ctggtcaacc aggttgcagg gtttcctgtc ctcacaggaa cgaagtccct aaagaaacag
     5461 tggcagccag gtttagcccc ggaattgact ggattccttt tttagggccc attggtatgg
     5521 ctttttcccc gtatcccccc aggtgtctgc aggctcaaag agcagcgaga agcgttcaga
     5581 ggaaagcgat cccgtgccac cttccccgtg cccgggctgt ccccgcacgc tgccggctcg
     5641 gggatgcggg gggagcgccg gaccggagcg gagccccggg cggctcgctg ctgcccccta
     5701 gcgggggagg gacgtaatta catccctggg ggctttgggg gggggctgtc cctgatatct
     5761 ataacaagaa aatatatata taataagtta tcacgtaagt agaacatgaa ataacaatat
     5821 aattatcgta tgagttaaat cttaaaagtc acgtaaaaga taatcatgcg tcattttgac
     5881 tcacgcggtc gttatagttc aaaatcagtg acacttaccg cattgacaag cacgcctcac
     5941 gggagctcca agcggcgact gagatgtcct aaatgcacag cgacggattc gcgctattta
     6001 gaaagagaga gcaatatttc aagaatgcat gcgtcaattt tacgcagact atctttctag
     6061 ggttaatcta gctgcatcag gatcatatcg tcgggtcttt tttccggctc agtcatcgcc
     6121 caagctggcg ctatctgggc atcggggagg aagaagcccg tgccttttcc cgcgaggttg
     6181 aagcggcatg gaaagagttt gccgaggatg actgctgctg cattgacgtt gagcgaaaac
     6241 gcacgtttac catgatgatt cgggaaggtg tggccatgca cgcctttaac ggtgaactgt
     6301 tcgttcaggc cacctgggat accagttcgt cgcggctttt ccggacacag ttccggatgg
     6361 tcagcccgaa gcgcatcagc aacccgaaca ataccggcga cagccggaac tgccgtgccg
     6421 gtgtgcagat taatgacagc ggtgcggcgc tgggatatta cgtcagcgag gacgggtatc
     6481 ctggctggat gccgcagaaa tggacatgga taccccgtga gttacccggc gggcgcgctt
     6541 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     6601 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     6661 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     6721 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     6781 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     6841 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     6901 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     6961 taggctccgc ccccctgacg agcatcacaa aaatcgacgc t
//
