LOCUS       sequence-323462-        4393 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  EYFP reporter vector for the ZF43x6-Spaced promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4393)
  AUTHORS   Donahue PS, Draut JW, Muldoon JJ, Edelstein HI, Bagheri N, Leonard 
            JN
  TITLE     The COMET toolkit for composing customizable genetic programs in 
            mammalian cells.
  JOURNAL   Nat Commun. 2020 Feb 7;11(1):779. doi: 10.1038/s41467-019-14147-5.
  PUBMED    32034124
REFERENCE   2  (bases 1 to 4393)
  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: "Nat 
            Commun."; date: "2020-02-7"; volume: "11(1):779. doi"; pages: " 
            10.1038/s41467-019-14147-5"
COMMENT     Sequence Label: pPD069 ZF43x6-Spaced_EYFP
FEATURES             Location/Qualifiers
     source          1..4393
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      284..293
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(290..292,293..295,296..1009)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(335..356)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(596..615)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     943..964
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1054..1071)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1060..1284
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1330..1758
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1417..1436)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1627..1648
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1781..1977
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      1828..1963
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1890..1909
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    2097..2218
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2134..2153)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2188..2207
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2267..2283)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2267..2283)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2280..2302)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2291..2307
                     /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(2315..2321,2322..2339,2340..2345))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2360..2381
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2498..2515)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2669..3257)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2749..2768)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3428..4219,4220..4288))
                     /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     4051..4070
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4289..4393)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cacatttccc cgaaaagtgc cacctgacgt cgacggatcg ggagatctga attcggatcc
       61 agagtgagga ctacctatag aagtaagagt gaggacaatc gtatacaagt agatccagag
      121 tgaggactac ctatagaagt aagagtgagg acaatcgtat acaagtagat ccagagtgag
      181 gactacctat agaagtaaga gtgaggacaa tcgtatacaa gtagatccct ctagagggta
      241 tataatgggg gccactagtc tactaccaga gctcatcgct agcgccacca tggtgagcaa
      301 gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa
      361 cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac
      421 cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac
      481 cttcggctac ggcctgcagt gcttcgcccg ctaccccgac cacatgaagc agcacgactt
      541 cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga
      601 cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat
      661 cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta
      721 caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt
      781 gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca
      841 gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcta
      901 ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt
      961 cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagtagg cggccgctcg
     1021 agtctagagg gcccgtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc
     1081 catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg
     1141 tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc
     1201 tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg
     1261 ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg
     1321 ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     1381 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct
     1441 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     1501 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac
     1561 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     1621 ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt
     1681 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     1741 aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttaggctcaa ttagtcagca
     1801 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     1861 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctctgcc
     1921 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag
     1981 ctcccgggag cttgtatatc cattttcgga ggttgggctt cggaatcgtt ttccgggacg
     2041 ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact
     2101 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     2161 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
     2221 atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc
     2281 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     2341 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     2401 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     2461 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     2521 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     2581 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     2641 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     2701 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     2761 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     2821 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     2881 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     2941 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     3001 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     3061 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     3121 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     3181 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     3241 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     3301 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     3361 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     3421 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     3481 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     3541 ctggccccag tgctgcaatg ataccgcgag atccacgctc accggctcca gatttatcag
     3601 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     3661 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     3721 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     3781 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     3841 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     3901 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     3961 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     4021 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     4081 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     4141 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     4201 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     4261 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     4321 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     4381 taggggttcc gcg
//
