LOCUS       sequence-323517-        4465 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  EYFP reporter vector for the ZF43x8-Spaced promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4465)
  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 4465)
  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: pPD089 ZF43x8-Spaced_EYFP
FEATURES             Location/Qualifiers
     source          1..4465
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      356..365
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(362..364,365..367,368..1081)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(407..428)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(668..687)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1015..1036
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1126..1143)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1132..1356
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1402..1830
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1489..1508)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1699..1720
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1853..2049
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      1900..2035
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1962..1981
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    2169..2290
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2206..2225)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2260..2279
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2339..2355)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2339..2355)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2352..2374)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2363..2379
                     /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(2387..2393,2394..2411,2412..2417))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2432..2453
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2570..2587)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2741..3329)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2821..2840)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3500..4291,4292..4360))
                     /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     4123..4142
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4361..4465)
                     /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 ccctctagag
      181 ggagatccag agtgaggact acctatagaa gtaagagtga ggacaatcgt atacaagtag
      241 atccagagtg aggactacct atagaagtaa gagtgaggac aatcgtatac aagtagatcc
      301 ctctagaggg tatataatgg gggccactag tctactacca gagctcatcg ctagcgccac
      361 catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga
      421 cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta
      481 cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac
      541 cctcgtgacc accttcggct acggcctgca gtgcttcgcc cgctaccccg accacatgaa
      601 gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt
      661 cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct
      721 ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca
      781 caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca agcagaagaa
      841 cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc
      901 cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca
      961 ctacctgagc taccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt
     1021 cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaagta
     1081 ggcggccgct cgagtctaga gggcccgttt aaacccgctg atcagcctcg actgtgcctt
     1141 ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg
     1201 ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt
     1261 gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca
     1321 atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct
     1381 ggggctctag ggggtatccc cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg
     1441 tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt
     1501 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc
     1561 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg
     1621 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg
     1681 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct
     1741 cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg
     1801 agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc agttaggctc
     1861 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
     1921 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
     1981 gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
     2041 ttttgcaaaa agctcccggg agcttgtata tccattttcg gaggttgggc ttcggaatcg
     2101 ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg
     2161 cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     2221 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     2281 atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt
     2341 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     2401 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
     2461 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     2521 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     2581 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     2641 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     2701 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     2761 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     2821 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     2881 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     2941 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     3001 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     3061 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     3121 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     3181 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     3241 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     3301 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     3361 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     3421 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     3481 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     3541 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     3601 agggcttacc atctggcccc agtgctgcaa tgataccgcg agatccacgc tcaccggctc
     3661 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     3721 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     3781 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     3841 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     3901 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     3961 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     4021 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     4081 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     4141 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     4201 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     4261 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     4321 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     4381 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     4441 aaaataaaca aataggggtt ccgcg
//
