LOCUS       sequence-320118-        4365 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Template for COMET promoters, expresses EYFP reporter upon promoter 
            activation..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4365)
  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 4365)
  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: pPD033 Reporter Template v1 (Far Spaced)
FEATURES             Location/Qualifiers
     source          1..4365
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      225..234
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(231..233,234..236,237..950)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(276..297)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(537..556)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     884..905
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(995..1012)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1001..1225
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1271..1699
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1358..1377)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1568..1589
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1722..1918
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      1769..1904
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1831..1850
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    2038..2159
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2075..2094)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2129..2148
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2208..2224)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2208..2224)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2221..2243)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2232..2248
                     /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(2256..2262,2263..2280,2281..2286))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2301..2322
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2439..2456)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2610..3198)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2690..2709)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3369..4160,4161..4229))
                     /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     3992..4011
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4230..4334)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctgaa ttcgagacca caggtctcgg atcctgagta gtgcgcgagc
       61 aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa tctgcttagg
      121 gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgatccc tctagagggt
      181 atataatggg ggccactagt ctactaccag agctcatcgc tagcgccacc atggtgagca
      241 agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa
      301 acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac ggcaagctga
      361 ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca
      421 ccttcggcta cggcctgcag tgcttcgccc gctaccccga ccacatgaag cagcacgact
      481 tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc ttcaaggacg
      541 acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg gtgaaccgca
      601 tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac aagctggagt
      661 acaactacaa cagccacaac gtctatatca tggccgacaa gcagaagaac ggcatcaagg
      721 tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc gaccactacc
      781 agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac tacctgagct
      841 accagtccgc cctgagcaaa gaccccaacg agaagcgcga tcacatggtc ctgctggagt
      901 tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagtag gcggccgctc
      961 gagtctagag ggcccgttta aacccgctga tcagcctcga ctgtgccttc tagttgccag
     1021 ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact
     1081 gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt
     1141 ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat
     1201 gctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg
     1261 gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     1321 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     1381 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     1441 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     1501 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     1561 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     1621 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     1681 caaaaattta acgcgaatta attctgtgga atgtgtgtca gttaggctca attagtcagc
     1741 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     1801 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc
     1861 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     1921 gctcccggga gcttgtatat ccattttcgg aggttgggct tcggaatcgt tttccgggac
     1981 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac
     2041 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     2101 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     2161 catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt
     2221 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     2281 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     2341 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     2401 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     2461 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     2521 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     2581 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     2641 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     2701 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     2761 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     2821 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     2881 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     2941 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     3001 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     3061 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     3121 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     3181 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     3241 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     3301 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     3361 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     3421 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     3481 tctggcccca gtgctgcaat gataccgcga gatccacgct caccggctcc agatttatca
     3541 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     3601 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     3661 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     3721 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     3781 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     3841 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     3901 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     3961 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     4021 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     4081 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     4141 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     4201 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     4261 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     4321 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtc
//
