LOCUS       sequence-274449-        7502 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Monomeric TagBFP2 and iCre-ERT2 recombinase (PAAAA N-terminus).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7502)
  AUTHORS   Nam HS, Capecchi MR
  TITLE     2A creER optimization plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7502)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pCAGGS-mTagBFP2-T2A-iCreERT2 (PAAAA)
FEATURES             Location/Qualifiers
     source          1..7502
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        11..390
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          670..1686
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1610..1632
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1694..1713
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1744..2454
                     /codon_start=1
                     /product="enhanced monomeric blue fluorescent protein 
                     (Subach et al., 2011)"
                     /label=mTagBFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTM
                     RIKVVEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGG
                     VLTATQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRND
                     MALKLVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAV
                     ARYCDLPSKLGHKLN"
     CDS             2476..2529
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             3577..4512
                     /codon_start=1
                     /product="mutated ligand-binding domain of the human 
                     estrogen receptor (Feil et al., 1997)"
                     /label=ERT2
                     /translation="AGDMRAANLWPSPLMIKRSKKNSLALSLTADQMVSALLDAEPPIL
                     YSEYDPTRPFSEASMMGLLTNLADRELVHMINWAKRVPGFVDLTLHDQVHLLECAWLEI
                     LMIGLVWRSMEHPVKLLFAPNLLLDRNQGKCVEGMVEIFDMLLATSSRFRMMNLQGEEF
                     VCLKSIILLNSGVYTFLSSTLKSLEEKDHIHRVLDKITDTLIHLMAKAGLTLQQQHQRL
                     AQLLLILSHIRHMSNKGMEHLYSMKCKNVVPLYDLLLEAADAHRLHAPTSRGGASVEET
                     DQSHLATAGSTSSHSLQKYYITGEAEGFPAT"
     CDS             3625..4518
                     /codon_start=1
                     /gene="ESR1"
                     /product="estrogen nuclear receptor alpha ligand-binding 
                     domain"
                     /label=ER-LBD
                     /translation="KRSKKNSLALSLTADQMVSALLDAEPPILYSEYDPTRPFSEASMM
                     GLLTNLADRELVHMINWAKRVPGFVDLTLHDQVHLLECAWLEILMIGLVWRSMEHPVKL
                     LFAPNLLLDRNQGKCVEGMVEIFDMLLATSSRFRMMNLQGEEFVCLKSIILLNSGVYTF
                     LSSTLKSLEEKDHIHRVLDKITDTLIHLMAKAGLTLQQQHQRLAQLLLILSHIRHMSNK
                     GMEHLYSMKCKNVVPLYDLLLEAADAHRLHAPTSRGGASVEETDQSHLATAGSTSSHSL
                     QKYYITGEAEGFPATA"
     primer_bind     complement(4530..4549)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4595..4650
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4649..4668)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(5011..5027)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5011..5027)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5024..5046)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5035..5051
                     /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(5059..5065,5066..5083,5084..5089))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5104..5125
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5184..5379
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5230..5365
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5385..5519
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5422..5441)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5476..5495
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5587..5604)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5758..6346)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5838..5857)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6517..7308,7309..7377))
                     /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     7140..7159
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7378..7482)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cacctgggtc gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta
       61 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
      121 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
      181 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
      241 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
      301 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
      361 atctacgtat tagtcatcgc tattaccatg gtcgaggtga gccccacgtt ctgcttcact
      421 ctccccatct cccccccctc cccaccccca attttgtatt tatttatttt ttaattattt
      481 tgtgcagcga tgggggcggg gggggggggg gggcgcgcgc cggggggggg gggggggggg
      541 gggggggggg ggggggsgrg gsggagaggt gcggcggcag ccaatcagag cggcgcgctc
      601 cgaaagtttc cttttatggc gaggcggcgg cggcggcggc cctataaaaa gcgaagcgcg
      661 cggcgggcgg gagtcgctgc gcgctgcctt cgccccgtgc cccgctccgc cgccgcctcg
      721 cgccgcccgc cccggctctg actgaccgcg ttactcccac aggtgagcgg gcgggacggc
      781 ccttctcctc cgggctgtaa ttagcgcttg gtttaatgac ggcttgtttc ttttctgtgg
      841 ctgcgtgaaa gccttgaggg gctccgggag ggccctttgt gcggggggag cggctcgggg
      901 ggtgcgtgcg tgtgtgtgtg cgtggggagc gccgcgtgcg gctccgcgct gcccggcggc
      961 tgtgagcgct gcgggcgcgg cgcggggctt tgtgcgctcc gcagtgtgcg cgaggggagc
     1021 gcggccgggg gcggtgcccc gcggtgcggg gggggctgcg aggggaacaa aggctgcgtg
     1081 cggggtgtgt gcgtgggggg gtgagcaggg ggtgtgggcg cgtcggtcgg gctgcaaccc
     1141 cccctgcacc cccctccccg agttgctgag cacggcccgg cttcgggtgc ggggctccgt
     1201 acggggcgtg gcgcggggct cgccgtgccg ggcggggggt ggcggcaggt gggggtgccg
     1261 ggcggggcgg ggccgcctcg ggccggggrg ggctcggggg rggggcgcgg cggcccccgg
     1321 agcgccggcg gctgtcgagg cgcggcgagc cgcagccatt gccttttatg gtaatcgtgc
     1381 gagagggcgc agggacttcc tttgtcccaa atctgtgcgg agccgaaatc tgggaggcgc
     1441 cgccgcaccc cctctagcgg gcgcggggcg aagcggtgcg gcgccggcag gaaggaaatg
     1501 ggcggggagg gccttcgtgc gtcgccgcgc cgccgtcccc ttctccctct ccagcctcgg
     1561 ggctgtccgc ggggggacgg ctgccttcgg gggggacggg gcagggcggg gttcggcttc
     1621 tggcgtgtga ccggcggctc tagagcctct gctaaccatg ttcatgcctt cttctttttc
     1681 ctacagctcc tgggcaacgt gctggttatt gtgctgtctc atcattttgg caaagaattc
     1741 gccatggtgt ctaagggcga agagctgatt aaggagaaca tgcacatgaa gctgtacatg
     1801 gagggcaccg tggacaacca tcacttcaag tgcacatccg agggcgaagg caagccctac
     1861 gagggcaccc agaccatgag aatcaaggtg gtcgagggcg gccctctccc cttcgccttc
     1921 gacatcctgg ctactagctt cctctacggc agcaagacct tcatcaacca cacccagggc
     1981 atccccgact tcttcaagca gtccttccct gagggcttca catgggagag agtcaccaca
     2041 tacgaagacg ggggcgtgct gaccgctacc caggacacca gcctccagga cggctgcctc
     2101 atctacaacg tcaagatcag aggggtgaac ttcacatcca acggccctgt gatgcagaag
     2161 aaaacactcg gctgggaggc cttcaccgag acgctgtacc ccgctgacgg cggcctggaa
     2221 ggcagaaacg acatggccct gaagctcgtg ggcgggagcc atctgatcgc aaacgccaag
     2281 accacatata gatccaagaa acccgctaag aacctcaaga tgcctggcgt ctactatgtg
     2341 gactacagac tggaaagaat caaggaggcc aacaacgaga cctacgtcga gcagcacgag
     2401 gtggcagtgg ccagatactg cgacctccct agcaaactgg ggcacaagct taatggcggc
     2461 tccggcggcc gctccgaggg ccgcggcagc ctgctgacat gcggcgacgt ggaagagaac
     2521 cctggccctg ccgctgcagc caacctgctg actgtgcacc aaaacctgcc tgccctccct
     2581 gtggatgcca cctctgatga agtcaggaag aacctgatgg acatgttcag ggacaggcag
     2641 gccttctctg aacacacctg gaagatgctc ctgtctgtgt gcagatcctg ggctgcctgg
     2701 tgcaagctga acaacaggaa atggttccct gctgaacctg aggatgtgag ggactacctc
     2761 ctgtacctgc aagccagagg cctggctgtg aagaccatcc aacagcacct gggccagctc
     2821 aacatgctgc acaggagatc tggcctgcct cgcccttctg actccaatgc tgtgtccctg
     2881 gtgatgagga gaatcagaaa ggagaatgtg gatgctgggg agagagccaa gcaggccctg
     2941 gcctttgaac gcactgactt tgaccaagtc agatccctga tggagaactc tgacagatgc
     3001 caggacatca ggaacctggc cttcctgggc attgcctaca acaccctgct gcgcattgcc
     3061 gaaattgcca gaatcagagt gaaggacatc tcccgcaccg atggtgggag aatgctgatc
     3121 cacattggca ggaccaagac cctggtgtcc acagctggtg tggagaaggc cctgtccctg
     3181 ggggttacca agctggtgga gagatggatc tctgtgtctg gtgtggctga tgaccccaac
     3241 aactacctgt tctgccgggt cagaaagaat ggtgtggctg ccccttctgc cacctcccaa
     3301 ctgtccaccc gggccctgga agggatcttt gaggccaccc accgcctgat ctatggtgcc
     3361 aaggatgact ctgggcagag atacctggcc tggtctggcc actctgccag agtgggtgct
     3421 gccagggaca tggccagggc tggtgtgtcc atccctgaaa tcatgcaggc tggtggctgg
     3481 accaatgtga acattgtgat gaactacatc agaaacctgg actctgagac tggggccatg
     3541 gtgaggctgc tagaggatgg ggacctcgag ccatctgctg gagacatgag agctgccaac
     3601 ctttggccaa gcccgctcat gatcaaacgc tctaagaaga acagcctggc cttgtccctg
     3661 acggccgacc agatggtcag tgccttgttg gatgctgagc cccccatact ctattccgag
     3721 tatgatccta ccagaccctt cagtgaagct tcgatgatgg gcttactgac caacctggca
     3781 gacagggagc tggttcacat gatcaactgg gcgaagaggg tgccaggctt tgtggatttg
     3841 accctccatg atcaggtcca ccttctagaa tgtgcctggc tagagatcct gatgattggt
     3901 ctcgtctggc gctccatgga gcacccagtg aagctactgt ttgctcctaa cttgctcttg
     3961 gacaggaacc agggaaaatg tgtagagggc atggtggaga tcttcgacat gctgctggct
     4021 acatcatctc ggttccgcat gatgaatctg cagggagagg agtttgtgtg cctcaaatct
     4081 attattttgc ttaattctgg agtgtacaca tttctgtcca gcaccctgaa gtctctggaa
     4141 gagaaggacc atatccaccg agtcctggac aagatcacag acactttgat ccacctgatg
     4201 gccaaggcag gcctgaccct gcagcagcag caccagcggc tggcccagct cctcctcatc
     4261 ctctcccaca tcaggcacat gagtaacaaa ggcatggagc atctgtacag catgaagtgc
     4321 aagaacgtgg tgcccctcta tgacctgctg ctggaggcgg cggacgccca ccgcctacat
     4381 gcgcccacta gccgtggagg ggcatccgtg gaggagacgg accaaagcca cttggccact
     4441 gcgggctcta cttcatcgca ttccttgcaa aagtattaca tcacggggga ggcagagggt
     4501 ttccctgcca cagcttgagg cgcgccggat ctttttccct ctgccaaaaa ttatggggac
     4561 atcatgaagc cccttgagca tctgacttct ggctaataaa ggaaatttat tttcattgca
     4621 atagtgtgtt ggaatttttt gtgtctctca ctcggaagga catatgggag ggcaaatcat
     4681 ttaaaacatc agaatgagta tttggtttag agtttggcaa catatgccca tatgctggct
     4741 gccatgaaca aaggttggct ataaagaggt catcagtata tgaaacagcc ccctgctgtc
     4801 cattccttat tccatagaaa agccttgact tgaggttaga ttttttttat attttgtttt
     4861 gtgttatttt tttctttaac atccctaaaa ttttccttac atgttttact agccagattt
     4921 ttcctcctct cctgactact cccagtcata gctgtccctc ttctcttatg gagatccctc
     4981 gacctgcagc ccaagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta
     5041 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     5101 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     5161 aaacctgtcg tgccagcgga tccgcatctc aattagtcag caaccatagt cccgccccta
     5221 actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga
     5281 ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag
     5341 tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctaacttg tttattgcag
     5401 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     5461 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctggatcc
     5521 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     5581 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     5641 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     5701 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     5761 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     5821 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     5881 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     5941 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     6001 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     6061 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     6121 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     6181 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     6241 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     6301 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     6361 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     6421 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     6481 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     6541 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     6601 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     6661 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     6721 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     6781 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     6841 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     6901 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     6961 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     7021 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     7081 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     7141 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     7201 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     7261 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     7321 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     7381 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     7441 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     7501 gc
//
