LOCUS       sequence-323529-        4604 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  EYFP reporter vector for the ZF43x10-Spaced-Mid promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4604)
  TITLE     Leonard Lab COMET plasmids
REFERENCE   2  (bases 1 to 4604)
  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"
COMMENT     Sequence Label: pPD138 ZF43x10-Spaced-Mid_EYFP
FEATURES             Location/Qualifiers
     source          1..4604
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      464..473
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(470..472,473..475,476..1189)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(515..536)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(776..795)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1123..1144
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1234..1251)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1240..1464
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1510..1938
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1597..1616)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1807..1828
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1961..2157
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      2008..2143
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2070..2089
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    2277..2398
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2314..2333)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2368..2387
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2447..2463)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2447..2463)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2460..2482)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2471..2487
                     /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(2495..2501,2502..2519,2520..2525))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2540..2561
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2678..2695)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2849..3437)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2929..2948)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3608..4399,4400..4468))
                     /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     4231..4250
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4469..4573)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctgaa ttcggatcca gagtgaggac tacctataga agtaagagtg
       61 aggacaatcg tatacaagta gatccagagt gaggactacc tatagaagta agagtgagga
      121 caatcgtata caagtagatc cagagtgagg actacctata gaagtaagag tgaggacaat
      181 cgtatacaag tagatccctc tagagggaga tccagagtga ggactaccta tagaagtaag
      241 agtgaggaca atcgtataca agtagatcca gagtgaggac tacctataga agtaagagtg
      301 aggacaatcg tatacaagta gatccctcta ggggagatcc catgaagaat ctgcttaggg
      361 ttaggcgttt tgcgctgctt cgcgatgtac gggccagata tacgatccct ctagagggta
      421 tataatgggg gccactagtc tactaccaga gctcatcgct agcgccacca tggtgagcaa
      481 gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa
      541 cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac
      601 cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac
      661 cttcggctac ggcctgcagt gcttcgcccg ctaccccgac cacatgaagc agcacgactt
      721 cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga
      781 cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat
      841 cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta
      901 caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt
      961 gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca
     1021 gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcta
     1081 ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt
     1141 cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagtagg cggccgctcg
     1201 agtctagagg gcccgtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc
     1261 catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg
     1321 tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc
     1381 tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg
     1441 ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg
     1501 ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     1561 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct
     1621 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     1681 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac
     1741 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     1801 ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt
     1861 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     1921 aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttaggctcaa ttagtcagca
     1981 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     2041 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctctgcc
     2101 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag
     2161 ctcccgggag cttgtatatc cattttcgga ggttgggctt cggaatcgtt ttccgggacg
     2221 ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact
     2281 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     2341 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
     2401 atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc
     2461 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     2521 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     2581 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     2641 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     2701 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     2761 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     2821 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     2881 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     2941 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     3001 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     3061 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     3121 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     3181 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     3241 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     3301 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     3361 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     3421 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     3481 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     3541 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     3601 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     3661 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     3721 ctggccccag tgctgcaatg ataccgcgag atccacgctc accggctcca gatttatcag
     3781 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     3841 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     3901 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     3961 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     4021 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     4081 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     4141 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     4201 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     4261 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     4321 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     4381 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     4441 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     4501 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     4561 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtc
//
