LOCUS       sequence-323877-        4422 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  EYFP reporter vector for the ZF43x2-Spaced-Far promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4422)
  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 4422)
  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: pPD144 ZF43x2-Spaced-Far_EYFP
FEATURES             Location/Qualifiers
     source          1..4422
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      282..291
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(288..290,291..293,294..1007)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(333..354)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(594..613)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     941..962
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1052..1069)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1058..1282
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1328..1756
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1415..1434)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1625..1646
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1779..1975
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      1826..1961
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1888..1907
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    2095..2216
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2132..2151)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2186..2205
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2265..2281)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2265..2281)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2278..2300)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2289..2305
                     /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(2313..2319,2320..2337,2338..2343))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2358..2379
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2496..2513)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2667..3255)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2747..2766)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3426..4217,4218..4286))
                     /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     4049..4068
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4287..4391)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctgaa ttcggatcca gagtgaggac tacctataga agtaagagtg
       61 aggacaatcg tatacaagta gatccctcta gagggagatc ctgagtagtg cgcgagcaaa
      121 atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt
      181 aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgatccctct agagggtata
      241 taatgggggc cactagtcta ctaccagagc tcatcgctag cgccaccatg gtgagcaagg
      301 gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg
      361 gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc
      421 tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccacct
      481 tcggctacgg cctgcagtgc ttcgcccgct accccgacca catgaagcag cacgacttct
      541 tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg
      601 gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg
      661 agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca
      721 actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc atcaaggtga
      781 acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac cactaccagc
      841 agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagctacc
      901 agtccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcg
      961 tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagtaggcg gccgctcgag
     1021 tctagagggc ccgtttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca
     1081 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc
     1141 ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg
     1201 gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct
     1261 ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg
     1321 tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     1381 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     1441 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     1501 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     1561 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     1621 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     1681 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     1741 aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt aggctcaatt agtcagcaac
     1801 catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc
     1861 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctctgcctc
     1921 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct
     1981 cccgggagct tgtatatcca ttttcggagg ttgggcttcg gaatcgtttt ccgggacgcc
     2041 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg
     2101 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     2161 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     2221 gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct
     2281 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     2341 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     2401 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     2461 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     2521 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     2581 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     2641 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     2701 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     2761 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     2821 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     2881 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     2941 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     3001 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     3061 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     3121 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     3181 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     3241 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     3301 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     3361 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     3421 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     3481 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     3541 ggccccagtg ctgcaatgat accgcgagat ccacgctcac cggctccaga tttatcagca
     3601 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     3661 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     3721 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     3781 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     3841 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     3901 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     3961 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     4021 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     4081 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     4141 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     4201 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     4261 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     4321 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     4381 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tc
//
