LOCUS       sequence-323878-        4478 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  EYFP reporter vector for the ZF43x4-Spaced-Far promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4478)
  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 4478)
  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: pPD145 ZF43x4-Spaced-Far_EYFP
FEATURES             Location/Qualifiers
     source          1..4478
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      338..347
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(344..346,347..349,350..1063)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(389..410)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(650..669)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     997..1018
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1108..1125)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1114..1338
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1384..1812
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1471..1490)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1681..1702
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1835..2031
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      1882..2017
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1944..1963
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    2151..2272
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2188..2207)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2242..2261
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2321..2337)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2321..2337)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2334..2356)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2345..2361
                     /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(2369..2375,2376..2393,2394..2399))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2414..2435
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2552..2569)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2723..3311)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2803..2822)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3482..4273,4274..4342))
                     /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     4105..4124
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4343..4447)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctgaa ttcggatcca gagtgaggac tacctataga agtaagagtg
       61 aggacaatcg tatacgagta gatccagagt gaggactacc tatagaagta agagtgagga
      121 caatcgtata caagtagatc cctctagagg gagatcctga gtagtgcgcg agcaaaattt
      181 aagctacaac aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt agggttaggc
      241 gttttgcgct gcttcgcgat gtacgggcca gatatacgat ccctctagag ggtatataat
      301 gggggccact agtctactac cagagctcat cgctagcgcc accatggtga gcaagggcga
      361 ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca
      421 caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa
      481 gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga ccaccttcgg
      541 ctacggcctg cagtgcttcg cccgctaccc cgaccacatg aagcagcacg acttcttcaa
      601 gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa
      661 ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct
      721 gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta
      781 caacagccac aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt
      841 caagatccgc cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa
      901 cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga gctaccagtc
      961 cgccctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac
     1021 cgccgccggg atcactctcg gcatggacga gctgtacaag taggcggccg ctcgagtcta
     1081 gagggcccgt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc cagccatctg
     1141 ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt
     1201 cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg
     1261 gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg catgctgggg
     1321 atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct agggggtatc
     1381 cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     1441 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
     1501 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     1561 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
     1621 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     1681 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
     1741 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     1801 ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggc tcaattagtc agcaaccata
     1861 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     1921 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag
     1981 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg
     2041 ggagcttgta tatccatttt cggaggttgg gcttcggaat cgttttccgg gacgccggct
     2101 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta
     2161 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     2221 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     2281 gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt
     2341 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     2401 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     2461 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     2521 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     2581 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     2641 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     2701 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     2761 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     2821 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     2881 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     2941 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     3001 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     3061 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     3121 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     3181 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     3241 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     3301 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     3361 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     3421 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     3481 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     3541 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     3601 ccagtgctgc aatgataccg cgagatccac gctcaccggc tccagattta tcagcaataa
     3661 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     3721 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     3781 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     3841 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     3901 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     3961 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     4021 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     4081 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     4141 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     4201 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     4261 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     4321 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     4381 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     4441 ttccgcgcac atttccccga aaagtgccac ctgacgtc
//
