LOCUS       sequence-319276-        7402 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Used for EGxxFP assay with CD55 intron 3, exon 4, and intron 4 
            sequences.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7402)
  AUTHORS   Hyodo T, Rahman ML, Karnan S, Ito T, Toyoda A, Ota A, Wahiduzzaman 
            M, Tsuzuki S, Okada Y, Hosokawa Y, Konishi H
  TITLE     Tandem Paired Nicking Promotes Precise Genome Editing with Scarce 
            Interference by p53.
  JOURNAL   Cell Rep. 2020 Jan 28;30(4):1195-1207.e7. doi: 
            10.1016/j.celrep.2019.12.064.
  PUBMED    31995758
REFERENCE   2  (bases 1 to 7402)
  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.celrep.2019.12"; journalName: "Cell Rep"; date: 
            "2020-01-28- 28"; volume: "30"; issue: "4"; pages: "1195-1207.e7"
COMMENT     Sequence Label: pCAG-EGxxFP-CD55
FEATURES             Location/Qualifiers
     source          1..7402
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        17..396
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        398..673
                     /label=chicken beta-actin promoter
     intron          674..1688
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1614..1636
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1696..1715
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1743..1752
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1749..1751,1752..1754,1755..2267)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIE"
     primer_bind     complement(1794..1815)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     3932..3953
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4082..4101)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4147..4202
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4201..4220)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4557..4576)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4558..4576)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4586..4603)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4586..4602)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4595..4617)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4744..5199
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4831..4850)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5041..5062
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5225..5329
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5330..5398,5399..6190)
                     /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     complement(5548..5567)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6361..6949
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6850..6869
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7103..7120
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7237..7258
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7273..7278,7279..7296,7297..7303)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7311..7327
                     /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)."
     primer_bind     7316..7338
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7335..7351
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7335..7351
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     7370..7390
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        7372..7390
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 gagctcatcg atactcgaca ttgattattg actagttatt aatagtaatc aattacgggg
       61 tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg
      121 cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata
      181 gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc
      241 cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac
      301 ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg
      361 cagtacatct acgtattagt catcgctatt accatggtcg aggtgagccc cacgttctgc
      421 ttcactctcc ccatctcccc cccctcccca cccccaattt tgtatttatt tattttttaa
      481 ttattttgtg cagcgatggg ggcggggggg gggggggsgc gcgccaggcg gggcggggcg
      541 gggcgagggg cggggcgggg cgaggcggag aggtgcggcg gcagccaatc agagcggcgc
      601 gctccgaaag tttcctttta tggcgaggcg gcggcggcgg cggccctata aaaagcgaag
      661 cgcgcggcgg gcgggagtcg ctgcgcgctg ccttcgcccc gtgccccgct ccgccgccgc
      721 ctcgcgccgc ccgccccggc tctgactgac cgcgttactc ccacaggtga gcgggcggga
      781 cggcccttct cctccgggct gtaattagcg cttggtttaa tgacggcttg tttcttttct
      841 gtggctgcgt gaaagccttg aggggctccg ggagggccct ttgtgcgggg ggagcggctc
      901 ggggggtgcg tgcgtgtgtg tgtgcgtggg gagcgccgcg tgcggctccg cgctgcccgg
      961 cggctgtgag cgctgcgggc gcggcgcggg gctttgtgcg ctccgcagtg tgcgcgaggg
     1021 gagcgcggcc gggggcggtg ccccgcggtg cggggggggc tgcgagggga acaaaggctg
     1081 cgtgcggggt gtgtgcgtgg gggggtgagc agggggtgtg ggcgcgtcgg tcgggctgca
     1141 accccccctg cacccccctc cccgagttgc tgagcacggc ccggcttcgg gtgcggggct
     1201 ccgtacgggg cgtggcgcgg ggctcgccgt gccgggcggg gggtggcggc aggtgggggt
     1261 gccgggcggg gcggggccgc ctcgggccgg ggagggctcg ggggaggggc gcggcggccc
     1321 ccggagcgcc ggcggctgtc gaggcgcggc gagccgcagc cattgccttt tatggtaatc
     1381 gtgcgagagg gcgcagggac ttcctttgtc ccaaatctgt gcggagccga aatctgggag
     1441 gcgccgccgc accccctcta gcgggcgcgg ggcgaagcgg tgcggcgccg gcaggaagga
     1501 aatgggcggg gagggccttc gtgcgtcgcc gcgccgccgt ccccttctcc ctctccagcc
     1561 tcggggctgt ccgcgggggg acggctgcct tcggggggga cggggcaggg cggggttcgg
     1621 cttctggcgt gtgaccggcg gctctagcct ctgctaacca tgttcatgcc ttcttctttt
     1681 tcctacagct cctgggcaac gtgctggtta ttgtgctgtc tcatcatttt ggcaaatcta
     1741 gagccgccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
     1801 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg
     1861 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
     1921 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc
     1981 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
     2041 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
     2101 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     2161 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc
     2221 agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc
     2281 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
     2341 acaaccactg aggatccgga atgtctttaa ctcatgctgt tttgtaacag cttgtttgca
     2401 tatagaaaaa ctcatgcaaa ttcattattt aggtcaacta aagattggtt tagtatttaa
     2461 catttgaaga tttctgcaag tatcatcatc tcaaattttt ttattaatca catttaaaat
     2521 tttatattga atcacatatt tactgaaatc agaatttaga gaattctggt ggtttaagaa
     2581 atcttccttt cttgccccca ttttactaaa tttaactgtg atgagctcat gatgaagggt
     2641 ctttgaggag agggattgtt ttaattgcta ctagctgtga ataccagtca tgaaactaac
     2701 aatctcacct taagtcctaa tttgttttct catttcctaa atacagggtt ttagtatgcc
     2761 caaacttacc ctgtgttaca tgagaaggag atggttgcac caaataatat gccacctggt
     2821 acatcaatct gaccatttcg tatttctccc ggattagggc atgatttctc tagaatagaa
     2881 tacaaattga ttttttaaaa ttaaattatc aggcatacac acaaagaagg taactatgtg
     2941 aggtggtaga tatgttaatt agcttgactg tgttaatcat ttcacaatgt gcacatatat
     3001 caaaacatca tgttgtacac ctaagataat atttgtcgat tatacctcaa taaagctcag
     3061 gtaaaaattt tgaaaaaatt atcaagcaca tatatgtaaa ttcatcatct gaaccattga
     3121 tatttcttat ccttagactt tttttttttt gagatggagt tttgctctgt tgcccaggct
     3181 ggagcacggt gacataatct cagctcactg caacctctgc ctcccaggtt caagggattc
     3241 tcctgcctca gccttccaag aagctgggat tacaggcatg cgccaccaca cccagctagt
     3301 ttttgtattt ttagtagagt cagggtttca tcatgttggc caggctactc tcaatctcct
     3361 gacctcaagt ggtgaattcg catatcgaat tcgatatcgg caagctgacc ctgaagttca
     3421 tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg
     3481 gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg
     3541 ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca
     3601 agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg
     3661 gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaaca
     3721 gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg aacttcaaga
     3781 tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc
     3841 ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccgccc
     3901 tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg
     3961 ccgggatcac tctcggcatg gacgagctgt acaagtaact cgagactcct caggtgcagg
     4021 ctgcctatca gaaggtggtg gctggtgtgg ccaatgccct ggctcacaaa taccactgag
     4081 atctttttcc ctctgccaaa aattatgggg acatcatgaa gccccttgag catctgactt
     4141 ctggctaata aaggaaattt attttcattg caatagtgtg ttggaatttt ttgtgtctct
     4201 cactcggaag gacatatggg agggcaaatc atttaaaaca tcagaatgag tatttggttt
     4261 agagtttggc aacatatgcc catatgctgg ctgccatgaa caaaggttgg ctataaagag
     4321 gtcatcagta tatgaaacag ccccctgctg tccattcctt attccataga aaagccttga
     4381 cttgaggtta gatttttttt atattttgtt ttgtgttatt tttttcttta acatccctaa
     4441 aattttcctt acatgtttta ctagccagat ttttcctcct ctcctgacta ctcccagtca
     4501 tagctgtccc tcttctctta tgaagatccc tcgacttaat taaggtaccc aattcgccct
     4561 atagtgagtc gtattacgcg cgctcactgg ccgtcgtttt acaacgtcgt gactgggaaa
     4621 accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc agctggcgta
     4681 atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg aatggcgaat
     4741 gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     4801 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
     4861 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     4921 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
     4981 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     5041 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
     5101 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     5161 ttaacgcgaa ttttaacaaa atattaacgc ttacaattta ggtggcactt ttcggggaaa
     5221 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     5281 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     5341 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     5401 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     5461 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     5521 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     5581 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     5641 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     5701 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     5761 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     5821 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     5881 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     5941 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     6001 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     6061 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     6121 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     6181 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     6241 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     6301 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     6361 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     6421 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     6481 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     6541 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     6601 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     6661 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     6721 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     6781 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     6841 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     6901 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     6961 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     7021 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     7081 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
     7141 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
     7201 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
     7261 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
     7321 ataacaattt cacacaggaa acagctatga ccatgattac gccaagcgcg caattaaccc
     7381 tcactaaagg gaacaaaagc tg
//
