LOCUS       sequence-272952-        6591 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  MADR donor plasmid for FlpO+Cre- mediated insertion of a SM_FP-FLAG,
            a cytoplasmic fluorescent protein which includes multiple FLAG 
            epitope tags.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6591)
  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 6591)
  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 smFP-FLAG (dark) WPRE
FEATURES             Location/Qualifiers
     source          1..6591
                     /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)."
     regulatory      1388..1397
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1397..1420
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1424..1447
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1451..1474
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2003..2026
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2033..2056
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2069..2092
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2099..2122
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2333..2356
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2360..2383
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2387..2410
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     intron          2459..2591
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     2574..2593
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    2600..3187
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2653..2673)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3070..3081)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3214..3231)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3220..3331
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3427..3447)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3427..3445)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3466..3482)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3584..3603)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3649..3704
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3703..3722)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     protein_bind    4042..4075
                     /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(4094..4110)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    4118..4134
                     /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(4142..4148,4149..4166,4167..4172))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4187..4208
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4267..4462
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4313..4448
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4468..4602
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4670..4687)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4841..5429)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4921..4940)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5600..6391,6392..6460))
                     /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     6223..6242
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6461..6565)
                     /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 ctagcgcaat
     1381 tgcggccgcc accatggact acaaggacga cgacgataaa ggagattaca aagatgacga
     1441 tgataagggc gattataagg acgacgatga caagggcggc gtgtcgaaag gcgaggagct
     1501 gttcacaggc gtggtcccca tcctggttga gctggacggt gacgttaacg gccacaaatt
     1561 tagcgtacgt ggtgagggcg agggtgacgc taccaacgga aagctgaccc tgaaattcat
     1621 ctgcacaact ggcaagctgc cggtgccctg gcccacgctg gtaaccactc tgggcggagg
     1681 cgttcaatgt ttcagtcgtt acccggatca catgaagcag catgacttct tcaagtcggc
     1741 catgcccgaa ggctacgtgc aggagcgcac catcagtttt aaggatgacg gcacctacaa
     1801 gactcgcgcc gaggtcaagt tcgaaggaga caccctggtt aatcgtatcg agctcaaagg
     1861 catcgatttc aaggaggacg gaaatattct gggccacaag ctggagtaca attttaacag
     1921 ccacaatgtt tacattactg ccgacaagca gaagaacggc atcaaggcta atttcaagat
     1981 tcgtcacaat gtggagggag gcgactataa ggatgacgat gacaaacaac aagactataa
     2041 ggacgatgat gacaaaggtc agcaaggcga ttacaaggac gatgacgata agcagcaaga
     2101 ttacaaggat gatgatgata agggaggcga cggaagcgtg cagctggcag atcattacca
     2161 acagaacacg cccattggtg acggtccagt cctgctgcct gataaccact acttgtccac
     2221 ccagtcggtg ctgtcgaagg acccgaacga gaagcgtgac cacatggtgc tgctggagtt
     2281 cgtgaccgcc gcaggaatca ccctcggcat ggatgagctt tacaagggcg gagattataa
     2341 agatgacgac gacaagggcg actacaaaga tgatgatgac aagggtgact acaaagacga
     2401 cgatgataaa taaaccggtg ctcgaggtac cgatgaatag ctaaggtcga ggccgcaggt
     2461 aagtatcaag gttacaagac aggtttaagg agaccaatag aaactgggct tgtcgagaca
     2521 gagaagactc ttgcgtttct gataggcacc tattggtctt actgacatcc actttgcctt
     2581 tctctccaca ggtgtcgaca atcaacctct ggattacaaa atttgtgaaa gattgactgg
     2641 tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta
     2701 tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct
     2761 gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt
     2821 tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc tttccgggac
     2881 tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg
     2941 ctggacaggg gctcggctgt tgggcactga cantccgtgg tgttgtcggg gaagctgacg
     3001 tcctttccat ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac gtccttctgc
     3061 tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg
     3121 cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc
     3181 tccccgcctg gaattcgagc tcggtacgat cagcctcgac tgtgccttct agttgccagc
     3241 catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg
     3301 tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc
     3361 tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat agcccagctt
     3421 ttgttccctt tagtgagggt taattgcgcg cttggcgtaa tcatggtcat agctgtttcc
     3481 tgtgtgaaat tgttatccgc taattcactc ctcaggtgca ggctgcctat cagaaggtgg
     3541 tggctggtgt ggccaatgcc ctggctcaca aataccactg agatcttttt ccctctgcca
     3601 aaaattatgg ggacatcatg aagccccttg agcatctgac ttctggctaa taaaggaaat
     3661 ttattttcat tgcaatagtg tgttggaatt ttttgtgtct ctcactcgga aggacatatg
     3721 ggagggcaaa tcatttaaaa catcagaatg agtatttggt ttagagtttg gcaacatatg
     3781 ccatatgctg gctgccatga acaaaggtgg ctataaagag gtcatcagta tatgaaacag
     3841 ccccctgctg tccattcctt attccataga aaagccttga cttgaggtta gatttttttt
     3901 atattttgtt ttgtgttatt tttttcttta acatccctaa aattttcctt acatgtttta
     3961 ctagccagat ttttcctcct ctcctgacta ctcccagtca tagctgtccc tcttctctta
     4021 tgaagatccc tcgacctgca ggaagttcct attctctaga aagtatagga acttcaagct
     4081 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     4141 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     4201 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     4261 ggatccgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     4321 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     4381 tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt
     4441 tggaggccta ggcttttgca aaaagctaac ttgtttattg cagcttataa tggttacaaa
     4501 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     4561 ggtttgtcca aactcatcaa tgtatcttat catgtctgga tccgctgcat taatgaatcg
     4621 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     4681 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     4741 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     4801 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     4861 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     4921 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     4981 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcaatgct
     5041 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     5101 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     5161 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     5221 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     5281 ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     5341 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     5401 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     5461 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     5521 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     5581 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     5641 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     5701 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     5761 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     5821 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     5881 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     5941 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     6001 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     6061 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     6121 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     6181 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     6241 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     6301 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     6361 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     6421 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     6481 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     6541 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct g
//
