LOCUS       sequence-257064-        7982 bp    DNA     circular SYN 14-APR-2025
DEFINITION  episomal expression of FlpO and Cre recombinases.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7982)
  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 7982)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 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: pCag FlpO-2A-Cre EV
FEATURES             Location/Qualifiers
     source          1..7982
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        50..429
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          707..1723
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1647..1669
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1731..1750
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     intron          1836..2065
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     2048..2067
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             3386..3439
                     /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             3440..4468
                     /codon_start=1
                     /product="site-specific recombinase"
                     /label=Cre
                     /note="Cre is a site-specific recombinase from 
                     bacteriophage P1. Recombination occurs at loxP sequences."
                     /translation="SNLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKMLL
                     SVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGLP
                     RPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAFL
                     GIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVER
                     WISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQR
                     YLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVRLLED
                     GD"
     polyA_signal    4611..4732
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4622..4641)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4676..4695
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(4947..4967)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4947..4965)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4986..5002)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4986..5002)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4999..5021)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5010..5026
                     /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(5034..5040,5041..5058,5059..5064))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5079..5100
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5217..5234)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5388..5976)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5468..5487)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6147..6938,6939..7007))
                     /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     6770..6789
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7008..7112)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(7138..7593)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7275..7296)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7487..7506
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7720..7742
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7734..7751
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7735..7751
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        7761..7779
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg acattgatta
       61 ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
      121 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
      181 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
      241 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
      301 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
      361 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
      421 attaccatgg tcgaggtgag ccccacgttc tgcttcactc tccccatctc ccccccctcc
      481 ccacccccaa ttttgtattt atttattttt taattatttt gtgcagcgat gggggcgggg
      541 gggggggggg ggcgcgccrg gsggggsggg gsggggsgrg gggsggggsg gggsgrggcg
      601 gagaggtgcg gcggcagcca atcagagcgg cgcgctccga aagtttcctt ttatggcgag
      661 gcggcggcgg cggcggccct ataaaaagcg aagcgcgcgg cgggcgggag tcgctgcgcg
      721 ctgccttcgc cccgtgcccc gctccgccgc cgcctcgcgc cgcccgcccc ggctctgact
      781 gaccgcgtta ctcccacagg tgagcgggcg ggacggccct tctcctccgg gctgtaatta
      841 gcgcttggtt taatgacggc ttgtttcttt tctgtggctg cgtgaaagcc ttgaggggct
      901 ccgggagggc cctttgtgcg gggggagcgg ctcggggggt gcgtgcgtgt gtgtgtgcgt
      961 ggggagcgcc gcgtgcggct ccgcgctgcc cggcggctgt gagcgctgcg ggcgcggcgc
     1021 ggggctttgt gcgctccgca gtgtgcgcga ggggagcgcg gccgggggcg gtgccccgcg
     1081 gtgcgggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg tgggggggtg
     1141 agcagggggt gtgggcgcgt cggtcgggct gcaacccccc ctgcaccccc ctccccgagt
     1201 tgctgagcac ggcccggctt cgggtgcggg gctccgtacg gggcgtggcg cggggctcgc
     1261 cgtgccgggc ggggggtggc ggcaggtggg ggtgccgggc ggggcggggc cgcctcgggc
     1321 cggggrgggc tcgggggagg ggcgcggcgg cccccggagc gccggcggct gtcgaggcgc
     1381 ggcgagccgc agccattgcc ttttatggta atcgtgcgag agggcgcagg gacttccttt
     1441 gtcccaaatc tgtgcggagc cgaaatctgg gaggcgccgc cgcaccccct ctagcgggcg
     1501 cggggcgaag cggtgcggcg ccggcaggaa ggaaatgggc ggggagggcc ttcgtgcgtc
     1561 gccgcgccgc cgtccccttc tccctctcca gcctcggggc tgtccgcggg gggacggctg
     1621 ccttcggggg ggacggggca gggcggggtt cggcttctgg cgtgtgaccg gcggctctag
     1681 agcctctgct aaccatgttc atgccttctt ctttttccta cagctcctgg gcaacgtgct
     1741 ggttattgtg ctgtctcatc attttggcaa atacgactca ctatagggag acccaagctt
     1801 ctagaattcg ctgtctgcga gggccagctg ttggggtgag tactccctct caaaagcggg
     1861 catgacttct gcgctaagat tgtcagtttc caaaaacgag gaggatttga tattcacctg
     1921 gcccgcggtg atgcctttga gggtggccgc gtccatctgg tcagaaaaga caatcttttt
     1981 gttgtcaagc ttgaggtgtg gcaggcttga gatctggcca tacacttgag tgacaatgac
     2041 atccactttg cctttctctc cacaggtgtc cactcccagg tccaactgca gcccaagcta
     2101 gaccatgagc cagttcgaca tcctgtgcaa gacccccccc aaggtgctgg tgcggcagtt
     2161 cgtggagaga ttcgagaggc ccagcggcga gaagatcgcc agctgtgccg ccgagctgac
     2221 ctacctgtgc tggatgatca cccacaacgg caccgccatc aagagggcca ccttcatgag
     2281 ctacaacacc atcatcagca acagcctgag cttcgacatc gtgaacaaga gcctgcagtt
     2341 caagtacaag acccagaagg ccaccatcct ggaggccagc ctgaagaagc tgatccccgc
     2401 ctgggagttc accatcatcc cttacaacgg ccagaagcac cagagcgaca tcaccgacat
     2461 cgtgtccagc ctgcagctgc agttcgagag cagcgaggag gccgacaagg gcaacagcca
     2521 cagcaagaag atgctgaagg ccctgctgtc cgagggcgag agcatctggg agatcaccga
     2581 gaagatcctg aacagcttcg agtacaccag caggttcacc aagaccaaga ccctgtacca
     2641 gttcctgttc ctggccacat tcatcaactg cggcaggttc agcgacatca agaacgtgga
     2701 ccccaagagc ttcaagctgg tgcagaacaa gtacctgggc gtgatcattc agtgcctggt
     2761 gaccgagacc aagacaagcg tgtccaggca catctacttt ttcagcgcca gaggcaggat
     2821 cgaccccctg gtgtacctgg acgagttcct gaggaacagc gagcccgtgc tgaagagagt
     2881 gaacaggacc ggcaacagca gcagcaacaa gcaggagtac cagctgctga aggacaacct
     2941 ggtgcgcagc tacaacaagg ccctgaagaa gaacgccccc taccccatct tcgctatcaa
     3001 gaacggccct aagagccaca tcggcaggca cctgatgacc agctttctga gcatgaaggg
     3061 cctgaccgag ctgacaaacg tggtgggcaa ctggagcgac aagagggcct ccgccgtggc
     3121 caggaccacc tacacccacc agatcaccgc catccccgac cactacttcg ccctggtgtc
     3181 caggtactac gcctacgacc ccatcagcaa ggagatgatc gccctgaagg acgagaccaa
     3241 ccccatcgag gagtggcagc acatcgagca gctgaagggc agcgccgagg gcagcatcag
     3301 ataccccgcc tggaacggca tcatcagcca ggaggtgctg gactacctga gcagctacat
     3361 caacaggcgg atctctagaa gagccgaggg caggggaagt cttctaacat gcggggacgt
     3421 ggaggaaaat cccgggccct ccaatttact gaccgtacac caaaatttgc ctgcattacc
     3481 ggtcgatgca acgagtgatg aggttcgcaa gaacctgatg gacatgttca gggatcgcca
     3541 ggcgttttct gagcatacct ggaaaatgct tctgtccgtt tgccggtcgt gggcggcatg
     3601 gtgcaagttg aataaccgga aatggtttcc cgcagaacct gaagatgttc gcgattatct
     3661 tctatatctt caggcgcgcg gtctggcagt aaaaactatc cagcaacatt tgggccagct
     3721 aaacatgctt catcgtcggt ccgggctgcc acgaccaagt gacagcaatg ctgtttcact
     3781 ggttatgcgg cgcattcgaa aagaaaacgt tgatgccggt gaacgtgcaa aacaggctct
     3841 agcgttcgaa cgcactgatt tcgaccaggt tcgttcactc atggaaaata gcgatcgctg
     3901 ccaggatata cgtaatctgg catttctggg gattgcttat aacaccctgt tacgtatagc
     3961 cgaaattgcc aggatcaggg ttaaagatat ctcacgtact gacggtggga gaatgttaat
     4021 ccatattggc agaacgaaaa cgctggttag caccgcaggt gtagagaagg cacttagcct
     4081 gggggtaact aaactggtcg agcgatggat ttccgtctct ggtgtagctg atgatccgaa
     4141 taactacctg ttttgccggg tcagaaaaaa tggtgttgcc gcgccatctg ccaccagcca
     4201 gctatcaact cgcgccctgg aagggatttt tgaagcaact catcgattga tttacggcgc
     4261 taaggatgac tctggtcaga gatacctggc ctggtctgga cacagtgccc gtgtcggagc
     4321 cgcgcgagat atggcccgcg ctggagtttc aataccggag atcatgcaag ctggtggctg
     4381 gaccaatgta aatattgtca tgaactatat ccgtaacctg gatagtgaaa caggggcaat
     4441 ggtgcgcctg ctggaagatg gcgattaagg atccgtacgc atttaagcat aaacacgcac
     4501 tatgccgttc ttctcatgta tatatatata caggcaacac gcagatatag gtgcgacgtg
     4561 aacagtgagc tgtatgtgcg ggtaccgagc tcctcgagga tcagacatga taagatacat
     4621 tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta tttgtgaaat
     4681 ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag ttatcctcga
     4741 ggagctcatg agcgcttgtt tcggcgtggg tatggtggca ggccccgtgg ccgggggact
     4801 gttgggcgcc atctccttgc atgcaccatt ccttgcggcg gcggtgctca acggcctcaa
     4861 cctactactg ggctgcttcc taatgcagga gtcgcataag ggagagcgtc gacctcgagg
     4921 gggggcccgg tacccagctt ttgttccctt tagtgagggt taattgcgcg cttggcgtaa
     4981 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
     5041 cgagccggga gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
     5101 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     5161 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     5221 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     5281 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     5341 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     5401 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     5461 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     5521 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     5581 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     5641 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     5701 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     5761 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     5821 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     5881 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     5941 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     6001 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     6061 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     6121 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     6181 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     6241 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     6301 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     6361 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     6421 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     6481 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     6541 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     6601 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     6661 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     6721 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     6781 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     6841 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     6901 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     6961 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     7021 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     7081 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctaaa
     7141 ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt
     7201 ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag
     7261 ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg
     7321 tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat
     7381 caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc
     7441 gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga
     7501 aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac
     7561 ccgccgcgct taatgcgccg ctacagggcg cgtcccattc gccattcagg ctgcgcaact
     7621 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat
     7681 gtgctgcaag gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa
     7741 cgacggccag tgagcgcgcg taatacgact cactataggg cgaattggag ctccaccgcg
     7801 gtctttcgga cttttgaaag tgatggtggt gggggaagga ttcgaacctt cgaagtcgat
     7861 gacggcagat ttagagtctg ctccctgctt gtgtgttgga ggtcgctgag tagtgcgcga
     7921 gcaaaattta agctacaaca aggcaaggct tgaccgacaa ttgcatgaag aatctgctta
     7981 gg
//
