LOCUS       sequence-323338-        4305 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  EYFP reporter vector for the ZF43x3-Spaced promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4305)
  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 4305)
  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: pPD063 ZF43x3-Spaced_EYFP
FEATURES             Location/Qualifiers
     source          1..4305
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      196..205
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(202..204,205..207,208..921)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(247..268)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(508..527)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     855..876
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(966..983)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    972..1196
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1242..1670
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1329..1348)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1539..1560
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1693..1889
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      1740..1875
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1802..1821
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    2009..2130
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2046..2065)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2100..2119
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2179..2195)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2179..2195)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2192..2214)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2203..2219
                     /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(2227..2233,2234..2251,2252..2257))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2272..2293
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2410..2427)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2581..3169)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2661..2680)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3340..4131,4132..4200))
                     /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     3963..3982
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4201..4305)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cacatttccc cgaaaagtgc cacctgacgt cgacggatcg ggagatctga attagagtga
       61 ggacaatcgt atacaagtag atccagagtg aggactacct atagaagtaa gagtgaggac
      121 aatcgtatac aagtagatcc ctctagaggg tatataatgg gggccactag tctactacca
      181 gagctcatcg ctagcgccac catggtgagc aagggcgagg agctgttcac cggggtggtg
      241 cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag
      301 ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag
      361 ctgcccgtgc cctggcccac cctcgtgacc accttcggct acggcctgca gtgcttcgcc
      421 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac
      481 gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg
      541 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag
      601 gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc
      661 atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag
      721 gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc
      781 gtgctgctgc ccgacaacca ctacctgagc taccagtccg ccctgagcaa agaccccaac
      841 gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc
      901 atggacgagc tgtacaagta ggcggccgct cgagtctaga gggcccgttt aaacccgctg
      961 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
     1021 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
     1081 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
     1141 gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc
     1201 tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc
     1261 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     1321 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     1381 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     1441 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     1501 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     1561 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
     1621 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg
     1681 aatgtgtgtc agttaggctc aattagtcag caaccatagt cccgccccta actccgccca
     1741 tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt
     1801 ttatttatgc agaggccgag gccgcctctg cctctgagct attccagaag tagtgaggag
     1861 gcttttttgg aggcctaggc ttttgcaaaa agctcccggg agcttgtata tccattttcg
     1921 gaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga
     1981 tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa
     2041 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     2101 tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta
     2161 gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat
     2221 tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
     2281 ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     2341 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     2401 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     2461 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     2521 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     2581 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     2641 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     2701 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     2761 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     2821 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     2881 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     2941 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     3001 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     3061 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     3121 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     3181 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     3241 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     3301 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     3361 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     3421 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     3481 agatccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     3541 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     3601 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     3661 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     3721 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     3781 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     3841 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     3901 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     3961 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     4021 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     4081 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     4141 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     4201 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     4261 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcg
//
