LOCUS       sequence-257066-        5912 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  MADR donor plasmid for FlpO+Cre-mediated insertion of a 
            TagBFP2-3XFlag cytoplasmic reporter protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5912)
  AUTHORS   Kim GB, Rincon Fernandez Pacheco D, Saxon D, Yang A, Sabet S, 
            Dutra-Clarke M, Levy R, Watkins A, Park H, Abbasi Akhtar A, Linesch 
            PW, Kobritz N, Chandra SS, Grausam K, Ayala-Sarmiento A, Molina J, 
            Sedivakova K, Hoang K, Tsyporin J, Gareau DS, Filbin MG, Bannykh S, 
            Santiskulvong C, Wang Y, Tang J, Suva ML, Chen B, Danielpour M, 
            Breunig JJ
  TITLE     Rapid Generation of Somatic Mouse Mosaics with Locus-Specific, 
            Stably Integrated Transgenic Elements.
  JOURNAL   Cell. 2019 Sep 19;179(1):251-267.e24. doi: 
            10.1016/j.cell.2019.08.013.
  PUBMED    31539496
REFERENCE   2  (bases 1 to 5912)
  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"; doi: 
            "10.1016/j.cell.2019.08"; journalName: "Cell"; date: "2019-09-19- 
            19"; volume: "179"; issue: "1"; pages: "251-267.e24"
COMMENT     Sequence Label: MADR pDonor TagBFP2-3XFlag (cyto) WPRE
FEATURES             Location/Qualifiers
     source          1..5912
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    32..493
                     /label=PGK poly(A) signal
                     /note="mouse phosphoglycerate kinase 1 polyadenylation 
                     signal"
     polyA_signal    641..775
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     polyA_signal    895..1029
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     polyA_signal    1149..1283
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     protein_bind    complement(1312..1345)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(1380..1396)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1381..1397)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     CDS             1434..2132
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV
                     VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK
                     LVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC
                     DLPSKLGHKLN"
     misc_feature    2230..2818
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2283..2303)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2701..2712)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(2903..2922)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2968..3023
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3022..3041)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     protein_bind    3363..3396
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     primer_bind     complement(3415..3431)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3415..3431)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3428..3450)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3439..3455
                     /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(3463..3469,3470..3487,3488..3493))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3508..3529
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3588..3783
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3634..3769
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3789..3923
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3991..4008)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4162..4750)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4242..4261)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4921..5712,5713..5781))
                     /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     5544..5563
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5782..5886)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtcgacatt gattattgac tagtggatcc gctgtaagtc tgcagaaatt gatgatctat
       61 taaacaataa agatgtccac taaaatggaa gtttttcctg tcatactttg ttaagaaggg
      121 tgagaacaga gtacctacat tttgaatgga aggattggag ctacgggggt gggggtgggg
      181 tgggattaga taaatgcctg ctctttactg aaggctcttt actattgctt tatgataatg
      241 tttcatagtt ggatatcata atttaaacaa gcaaaaccaa attaagggcc agctcattcc
      301 tcccactcat gatctataga tctatagatc tctcgtggga tcattgtttt tctcttgatt
      361 cccactttgt ggttctaagt actgtggttt ccaaatgtgt cagtttcata gcctgaagaa
      421 cgagatcagc agcctctgtt ccacatacac ttcattctca gtattgtttt gccaagttct
      481 aattccatca gaagcttgca gatctgcgac tctagaggat ctgcgactct agaggatcat
      541 aatcagccat accacatttg tagaggtttt acttgcttta aaaaacctcc cacacctccc
      601 cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt aacttgttta ttgcagctta
      661 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
      721 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct ggatctgcga
      781 ctctagagga tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac
      841 ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg
      901 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
      961 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     1021 gtctggatct gcgactctag aggatcataa tcagccatac cacatttgta gaggttttac
     1081 ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg
     1141 ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     1201 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     1261 atgtatctta tcatgtctgg atccccatca agctgatccg gaacccttaa tataacttcg
     1321 tatagcatac attatacgaa gttattaggt ccctcgacct gcagcccaag ctagcttatc
     1381 gataccgtcg acctcgaggg ggggaattct gcagtcgagg taccgccgcc accatgagcg
     1441 agctgattaa ggagaacatg cacatgaagc tgtacatgga gggcaccgtg gacaaccatc
     1501 acttcaagtg cacatccgag ggcgaaggca agccctacga gggcacccag accatgagaa
     1561 tcaaggtggt cgagggcggc cctctcccct tcgccttcga catcctggct actagcttcc
     1621 tctacggcag caagaccttc atcaaccaca cccagggcat ccccgacttc ttcaagcagt
     1681 ccttccctga gggcttcaca tgggagagag tcaccacata cgaagacggg ggcgtgctga
     1741 ccgctaccca ggacaccagc ctccaggacg gctgcctcat ctacaacgtc aagatcagag
     1801 gggtgaactt cacatccaac ggccctgtga tgcagaagaa aacactcggc tgggaggcct
     1861 tcaccgagac gctgtacccc gctgacggcg gcctggaagg cagaaacgac atggccctga
     1921 agctcgtggg cgggagccat ctgatcgcaa acgccaagac cacatataga tccaagaaac
     1981 ccgctaagaa cctcaagatg cctggcgtct actatgtgga ctacagactg gaaagaatca
     2041 aggaggccaa caacgagacc tacgtcgagc agcacgaggt ggcagtggcc agatactgcg
     2101 acctccctag caaactgggg cacaagctta attccggact cgactacaaa gaccatgacg
     2161 gtgattataa agatcatgac atcgactaca aggatgacga tgactaagcg gccgcactcc
     2221 tcaggtgcta atcaacctct ggattacaaa atttgtgaaa gattgactgg tattcttaac
     2281 tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta tcatgctatt
     2341 gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct gtctctttat
     2401 gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca
     2461 acccccactg gttggggcat tgccaccacc tgtcagctcc tttccgggac tttcgctttc
     2521 cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg
     2581 gctcggctgt tgggcactga caattccgtg gtgttgtcgg ggaaatcatc gtcctttcct
     2641 tggctgctcg cctgtgttgc cacctggatt ctgcgcggga cgtccttctg ctacgtccct
     2701 tcggccctca atccagcgga ccttccttcc cgcggcctgc tgccggctct gcggcctctt
     2761 ccgcgtcttc gccttcgccc tcagacgagt cggatctccc tttgggccgc ctccccgcag
     2821 gccgcactcc tcaggtgcag gctgcctatc agaaggtggt ggctggtgtg gccaatgccc
     2881 tggctcacaa ataccactga gatctttttc cctctgccaa aaattatggg gacatcatga
     2941 agccccttga gcatctgact tctggctaat aaaggaaatt tattttcatt gcaatagtgt
     3001 gttggaattt tttgtgtctc tcactcggaa ggacatatgg gagggcaaat catttaaaac
     3061 atcagaatga gtatttggtt tagagtttgg caacatatgc ccatatgctg gctgccatga
     3121 acaaaggttg gctataaaga ggtcatcagt atatgaaaca gccccctgct gtccattcct
     3181 tattccatag aaaagccttg acttgaggtt agattttttt tatattttgt tttgtgttat
     3241 ttttttcttt aacatcccta aaattttcct tacatgtttt actagccaga tttttcctcc
     3301 tctcctgact actcccagtc atagctgtcc ctcttctctt atggagatcc ctcgacctgc
     3361 aggaagttcc tattctctag aaagtatagg aacttcaagc ttggcgtaat catggtcata
     3421 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     3481 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     3541 ctcactgccc gctttccagt cgggaaacct gtcgtgccag cggatccgca tctcaattag
     3601 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
     3661 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     3721 tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     3781 aaaaagctaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     3841 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     3901 atgtatctta tcatgtctgg atccgctgca ttaatgaatc ggccaacgcg cggggagagg
     3961 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     4021 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     4081 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     4141 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     4201 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     4261 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     4321 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     4381 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     4441 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     4501 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     4561 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     4621 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     4681 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     4741 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     4801 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     4861 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     4921 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     4981 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     5041 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     5101 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     5161 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     5221 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     5281 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     5341 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     5401 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     5461 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     5521 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     5581 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     5641 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     5701 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     5761 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     5821 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     5881 tccgcgcaca tttccccgaa aagtgccacc tg
//
