LOCUS       sequence-323519-        4521 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  EYFP reporter vector for the ZF43x10-Spaced promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4521)
  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 4521)
  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: pPD091 ZF43x10-Spaced_EYFP
FEATURES             Location/Qualifiers
     source          1..4521
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      412..421
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(418..420,421..423,424..1137)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(463..484)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(724..743)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1071..1092
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1182..1199)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1188..1412
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1458..1886
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1545..1564)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1755..1776
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1909..2105
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      1956..2091
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2018..2037
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    2225..2346
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2262..2281)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2316..2335
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2395..2411)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2395..2411)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2408..2430)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2419..2435
                     /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(2443..2449,2450..2467,2468..2473))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2488..2509
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2626..2643)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2797..3385)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2877..2896)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3556..4347,4348..4416))
                     /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     4179..4198
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4417..4521)
                     /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 ctagagggag
      241 atccagagtg aggactacct atagaagtaa gagtgaggac aatcgtatac aagtagatcc
      301 agagtgagga ctacctatag aagtaagagt gaggacaatc gtatacaagt agatccctct
      361 agagggtata taatgggggc cactagtcta ctaccagagc tcatcgctag cgccaccatg
      421 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
      481 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc
      541 aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc
      601 gtgaccacct tcggctacgg cctgcagtgc ttcgcccgct accccgacca catgaagcag
      661 cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc
      721 aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg
      781 aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
      841 ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc
      901 atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac
      961 cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac
     1021 ctgagctacc agtccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg
     1081 ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagtaggcg
     1141 gccgctcgag tctagagggc ccgtttaaac ccgctgatca gcctcgactg tgccttctag
     1201 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
     1261 tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca
     1321 ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag
     1381 caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg
     1441 ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     1501 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt
     1561 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
     1621 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga
     1681 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     1741 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt
     1801 ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct
     1861 gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt aggctcaatt
     1921 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     1981 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     2041 cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     2101 gcaaaaagct cccgggagct tgtatatcca ttttcggagg ttgggcttcg gaatcgtttt
     2161 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     2221 ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     2281 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     2341 atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata
     2401 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     2461 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     2521 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     2581 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     2641 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     2701 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     2761 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     2821 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     2881 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     2941 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     3001 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     3061 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     3121 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     3181 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     3241 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     3301 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     3361 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     3421 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     3481 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     3541 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     3601 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     3661 cttaccatct ggccccagtg ctgcaatgat accgcgagat ccacgctcac cggctccaga
     3721 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     3781 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     3841 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     3901 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     3961 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     4021 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     4081 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     4141 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     4201 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     4261 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     4321 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     4381 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     4441 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     4501 taaacaaata ggggttccgc g
//
