LOCUS       sequence-320117-        4308 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 4308)
  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 4308)
  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: pPD032 Reporter Template v1 (Mid Spaced)
FEATURES             Location/Qualifiers
     source          1..4308
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      168..177
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(174..176,177..179,180..893)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(219..240)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(480..499)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     827..848
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(938..955)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    944..1168
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1214..1642
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1301..1320)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1511..1532
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1665..1861
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      1712..1847
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1774..1793
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    1981..2102
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2018..2037)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2072..2091
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2151..2167)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2151..2167)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2164..2186)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2175..2191
                     /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(2199..2205,2206..2223,2224..2229))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2244..2265
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2382..2399)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2553..3141)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2633..2652)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3312..4103,4104..4172))
                     /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     3935..3954
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4173..4277)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctgaa ttcgagacca caggtctcgg atcccatgaa gaatctgctt
       61 agggttaggc gttttgcgct gcttcgcgat gtacgggcca gatatacgat ccctctagag
      121 ggtatataat gggggccact agtctactac cagagctcat cgctagcgcc accatggtga
      181 gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg
      241 taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc
      301 tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga
      361 ccaccttcgg ctacggcctg cagtgcttcg cccgctaccc cgaccacatg aagcagcacg
      421 acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg
      481 acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc
      541 gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg
      601 agtacaacta caacagccac aacgtctata tcatggccga caagcagaag aacggcatca
      661 aggtgaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc gccgaccact
      721 accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga
      781 gctaccagtc cgccctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg
      841 agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag taggcggccg
      901 ctcgagtcta gagggcccgt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc
      961 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     1021 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     1081 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
     1141 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct
     1201 agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     1261 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     1321 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     1381 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
     1441 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     1501 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
     1561 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
     1621 taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggc tcaattagtc
     1681 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     1741 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     1801 tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     1861 aaagctcccg ggagcttgta tatccatttt cggaggttgg gcttcggaat cgttttccgg
     1921 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc
     1981 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     2041 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     2101 tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg
     2161 tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata
     2221 aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca
     2281 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc
     2341 gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg
     2401 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
     2461 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
     2521 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     2581 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     2641 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     2701 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     2761 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     2821 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     2881 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     2941 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta
     3001 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     3061 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     3121 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     3181 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     3241 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     3301 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     3361 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     3421 ccatctggcc ccagtgctgc aatgataccg cgagatccac gctcaccggc tccagattta
     3481 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     3541 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     3601 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt
     3661 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     3721 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     3781 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     3841 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     3901 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact
     3961 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     4021 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     4081 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     4141 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc
     4201 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa
     4261 caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtc
//
