LOCUS       sequence-274459-        7505 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Monomeric TagBFP2 and iCre-ERT2 recombinase (PAGSAS N-terminus).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7505)
  AUTHORS   Nam HS, Capecchi MR
  TITLE     2A creER optimization plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7505)
  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            "Unpublished"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 (PAGSAS)
FEATURES             Location/Qualifiers
     source          1..7505
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        11..390
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        392..669
                     /label=chicken beta-actin promoter
     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             3580..4515
                     /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             3628..4521
                     /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(4533..4552)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4598..4653
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4652..4671)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(5014..5030)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5014..5030)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5027..5049)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5038..5054
                     /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(5062..5068,5069..5086,5087..5092))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5107..5128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5187..5382
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5233..5368
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5388..5522
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5425..5444)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5479..5498
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5590..5607)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5761..6349)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5841..5860)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6520..7311,7312..7380))
                     /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     7143..7162
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7381..7485)
                     /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 gsggggsgrg gcggagaggt 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 ggccgccycg 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 ccggcagcgc cagcaacctg ctgactgtgc accaaaacct gcctgccctc
     2581 cctgtggatg ccacctctga tgaagtcagg aagaacctga tggacatgtt cagggacagg
     2641 caggccttct ctgaacacac ctggaagatg ctcctgtctg tgtgcagatc ctgggctgcc
     2701 tggtgcaagc tgaacaacag gaaatggttc cctgctgaac ctgaggatgt gagggactac
     2761 ctcctgtacc tgcaagccag aggcctggct gtgaagacca tccaacagca cctgggccag
     2821 ctcaacatgc tgcacaggag atctggcctg cctcgccctt ctgactccaa tgctgtgtcc
     2881 ctggtgatga ggagaatcag aaaggagaat gtggatgctg gggagagagc caagcaggcc
     2941 ctggcctttg aacgcactga ctttgaccaa gtcagatccc tgatggagaa ctctgacaga
     3001 tgccaggaca tcaggaacct ggccttcctg ggcattgcct acaacaccct gctgcgcatt
     3061 gccgaaattg ccagaatcag agtgaaggac atctcccgca ccgatggtgg gagaatgctg
     3121 atccacattg gcaggaccaa gaccctggtg tccacagctg gtgtggagaa ggccctgtcc
     3181 ctgggggtta ccaagctggt ggagagatgg atctctgtgt ctggtgtggc tgatgacccc
     3241 aacaactacc tgttctgccg ggtcagaaag aatggtgtgg ctgccccttc tgccacctcc
     3301 caactgtcca cccgggccct ggaagggatc tttgaggcca cccaccgcct gatctatggt
     3361 gccaaggatg actctgggca gagatacctg gcctggtctg gccactctgc cagagtgggt
     3421 gctgccaggg acatggccag ggctggtgtg tccatccctg aaatcatgca ggctggtggc
     3481 tggaccaatg tgaacattgt gatgaactac atcagaaacc tggactctga gactggggcc
     3541 atggtgaggc tgctagagga tggggacctc gagccatctg ctggagacat gagagctgcc
     3601 aacctttggc caagcccgct catgatcaaa cgctctaaga agaacagcct ggccttgtcc
     3661 ctgacggccg accagatggt cagtgccttg ttggatgctg agccccccat actctattcc
     3721 gagtatgatc ctaccagacc cttcagtgaa gcttcgatga tgggcttact gaccaacctg
     3781 gcagacaggg agctggttca catgatcaac tgggcgaaga gggtgccagg ctttgtggat
     3841 ttgaccctcc atgatcaggt ccaccttcta gaatgtgcct ggctagagat cctgatgatt
     3901 ggtctcgtct ggcgctccat ggagcaccca gtgaagctac tgtttgctcc taacttgctc
     3961 ttggacagga accagggaaa atgtgtagag ggcatggtgg agatcttcga catgctgctg
     4021 gctacatcat ctcggttccg catgatgaat ctgcagggag aggagtttgt gtgcctcaaa
     4081 tctattattt tgcttaattc tggagtgtac acatttctgt ccagcaccct gaagtctctg
     4141 gaagagaagg accatatcca ccgagtcctg gacaagatca cagacacttt gatccacctg
     4201 atggccaagg caggcctgac cctgcagcag cagcaccagc ggctggccca gctcctcctc
     4261 atcctctccc acatcaggca catgagtaac aaaggcatgg agcatctgta cagcatgaag
     4321 tgcaagaacg tggtgcccct ctatgacctg ctgctggagg cggcggacgc ccaccgccta
     4381 catgcgccca ctagccgtgg aggggcatcc gtggaggaga cggaccaaag ccacttggcc
     4441 actgcgggct ctacttcatc gcattccttg caaaagtatt acatcacggg ggaggcagag
     4501 ggtttccctg ccacagcttg aggcgcgccg gatctttttc cctctgccaa aaattatggg
     4561 gacatcatga agccccttga gcatctgact tctggctaat aaaggaaatt tattttcatt
     4621 gcaatagtgt gttggaattt tttgtgtctc tcactcggaa ggacatatgg gagggcaaat
     4681 catttaaaac atcagaatga gtatttggtt tagagtttgg caacatatgc ccatatgctg
     4741 gctgccatga acaaaggttg gctataaaga ggtcatcagt atatgaaaca gccccctgct
     4801 gtccattcct tattccatag aaaagccttg acttgaggtt agattttttt tatattttgt
     4861 tttgtgttat ttttttcttt aacatcccta aaattttcct tacatgtttt actagccaga
     4921 tttttcctcc tctcctgact actcccagtc atagctgtcc ctcttctctt atggagatcc
     4981 ctcgacctgc agcccaagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg
     5041 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
     5101 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
     5161 gggaaacctg tcgtgccagc ggatccgcat ctcaattagt cagcaaccat agtcccgccc
     5221 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     5281 tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag
     5341 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctaac ttgtttattg
     5401 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     5461 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgga
     5521 tccgctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     5581 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     5641 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     5701 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     5761 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     5821 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     5881 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     5941 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     6001 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     6061 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     6121 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     6181 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     6241 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     6301 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     6361 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     6421 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     6481 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     6541 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     6601 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     6661 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     6721 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     6781 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     6841 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     6901 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     6961 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     7021 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     7081 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     7141 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     7201 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     7261 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     7321 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     7381 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     7441 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     7501 agtgc
//
