LOCUS       sequence-378927-        7899 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  coexpression of Nanoluc, copGFP and puromycin resistant marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7899)
  TITLE     Oka Lab Plasmids 2022
REFERENCE   2  (bases 1 to 7899)
  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: pCDH-CMV5-N-cG-P
FEATURES             Location/Qualifiers
     source          1..7899
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        6..232
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             233..413
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    460..585
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1078..1311
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1496..1540
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    1807..1923
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2013..2216
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2167..2187
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2213..2237
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             2301..2813
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     CDS             2829..2885
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2910..3572
                     /codon_start=1
                     /product="green fluorescent protein 2 from Pontellina 
                     plumata, also known as ppluGFP2 (Shagin et al., 2004)"
                     /label=CopGFP
                     /translation="PAMEIECRITGTLNGVEFELVGGGEGTPKQGRMTNKMKSTKGALT
                     FSPYLLSHVMGYGFYHFGTYPSGYENPFLHAINNGGYTNTRIEKYEDGGVLHVSFSYRY
                     EAGRVIGDFKVVGTGFPEDSVIFTDKIIRSNATVEHLHPMGDNVLVGSFARTFSLRDGG
                     YYSFVVDSHMHFKSAIHPSILQNGGPMFAFRRVEELHSNTELGIVEYQHAFKTPIAFA"
     CDS             3657..3710
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             3711..4310
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3711..3730)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4207..4227
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(4376..4396)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     LTR             4640..4873
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4945..5066
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4982..5001)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5036..5055
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      5106..5241
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5274..5290)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5274..5290)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5287..5309)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5298..5314
                     /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(5322..5328,5329..5346,5347..5352))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5367..5388
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5505..5522)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5676..6264)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5756..5775)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6435..7226,7227..7295))
                     /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     7058..7077
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7296..7400)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7468..7486
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7524..7546)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7646..7665
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7859..7881
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7873..7890
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7874..7890
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 acgcgtgtag tcttatgcaa tactcttgta gtcttgcaac atggtaacga tgagttagca
       61 acatgcctta caaggagaga aaaagcaccg tgcatgccga ttggtggaag taaggtggta
      121 cgatcgtgcc ttattaggaa ggcaacagac gggtctgaca tggattggac gaaccactga
      181 attgccgcat tgcagagata ttgtatttaa gtgcctagct cgatacataa acgggtctct
      241 ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc cactgcttaa
      301 gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc
      361 tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta gcagtggcgc
      421 ccgaacaggg acttgaaagc gaaagggaaa ccagaggagc tctctcgacg caggactcgg
      481 cttgctgaag cgcgcacggc aagaggcgag gggcggcgac tggtgagtac gccaaaaatt
      541 ttgactagcg gaggctagaa ggagagagat gggtgcgaga gcgtcagtat taagcggggg
      601 agaattagat cgcgatggga aaaaattcgg ttaaggccag ggggaaagaa aaaatataaa
      661 ttaaaacata tagtatgggc aagcagggag ctagaacgat tcgcagttaa tcctggcctg
      721 ttagaaacat cagaaggctg tagacaaata ctgggacagc tacaaccatc ccttcagaca
      781 ggatcagaag aacttagatc attatataat acagtagcaa ccctctattg tgtgcatcaa
      841 aggatagaga taaaagacac caaggaagct ttagacaaga tagaggaaga gcaaaacaaa
      901 agtaagacca ccgcacagca agcggccact gatcttcaga cctggaggag gagatatgag
      961 ggacaattgg agaagtgaat tatataaata taaagtagta aaaattgaac cattaggagt
     1021 agcacccacc aaggcaaaga gaagagtggt gcagagagaa aaaagagcag tgggaatagg
     1081 agctttgttc cttgggttct tgggagcagc aggaagcact atgggcgcag cgtcaatgac
     1141 gctgacggta caggccagac aattattgtc tggtatagtg cagcagcaga acaatttgct
     1201 gagggctatt gaggcgcaac agcatctgtt gcaactcaca gtctggggca tcaagcagct
     1261 ccaggcaaga atcctggctg tggaaagata cctaaaggat caacagctcc tggggatttg
     1321 gggttgctct ggaaaactca tttgcaccac tgctgtgcct tggaatgcta gttggagtaa
     1381 taaatctctg gaacagattt ggaatcacac gacctggatg gagtgggaca gagaaattaa
     1441 caattacaca agcttaatac actccttaat tgaagaatcg caaaaccagc aagaaaagaa
     1501 tgaacaagaa ttattggaat tagataaatg ggcaagtttg tggaattggt ttaacataac
     1561 aaattggctg tggtatataa aattattcat aatgatagta ggaggcttgg taggtttaag
     1621 aatagttttt gctgtacttt ctatagtgaa tagagttagg cagggatatt caccattatc
     1681 gtttcagacc cacctcccaa ccccgagggg acccgacagg cccgaaggaa tagaagaaga
     1741 aggtggagag agagacagag acagatccat tcgattagtg aacggatctc gacggtatcg
     1801 gttaactttt aaaagaaaag gggggattgg ggggtacagt gcaggggaaa gaatagtaga
     1861 cataatagca acagacatac aaactaaaga attacaaaaa caaattacaa aattcaaaat
     1921 tttatcgata ctagtattat gcccagtaca tgaccttatg ggactttcct acttggcagt
     1981 acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg
     2041 ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg
     2101 ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc
     2161 cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ttatataagc agagctcgtt
     2221 tagtgaaccg tcagatcgcc tggagacgcc atccacgctg ttttgacctc catagaagat
     2281 tctagagcta gggatccacc atggtcttca cactcgaaga tttcgttggg gactggcgac
     2341 agacagccgg ctacaacctg gaccaagtcc ttgaacaggg aggtgtgtcc agtttgtttc
     2401 agaatctcgg ggtgtccgta actccgatcc aaaggattgt cctgagcggt gaaaatgggc
     2461 tgaagatcga catccatgtc atcatcccgt atgaaggtct gagcggcgac caaatgggcc
     2521 agatcgaaaa aatttttaag gtggtgtacc ctgtggatga tcatcacttt aaggtgatcc
     2581 tgcactatgg cacactggta atcgacgggg ttacgccgaa catgatcgac tatttcggac
     2641 ggccgtatga aggcatcgcc gtgttcgacg gcaaaaagat cactgtaaca gggaccctgt
     2701 ggaacggcaa caaaattatc gacgagcgcc tgatcaaccc cgacggctcc ctgctgttcc
     2761 gagtaaccat caacggagtg accggctggc ggctgtgcga acgcattctg gcgaagcttg
     2821 gaagcggagc tactaacttc agcctgctga agcaggctgg agacgtggag gagaaccctg
     2881 gacctatgga gagcgacgag agcggcctgc ccgccatgga gatcgagtgc cgcatcaccg
     2941 gcaccctgaa cggcgtggag ttcgagctgg tgggcggcgg agagggcacc cccaagcagg
     3001 gccgcatgac caacaagatg aagagcacca aaggcgccct gaccttcagc ccctacctgc
     3061 tgagccacgt gatgggctac ggcttctacc acttcggcac ctaccccagc ggctacgaga
     3121 accccttcct gcacgccatc aacaacggcg gctacaccaa cacccgcatc gagaagtacg
     3181 aggacggcgg cgtgctgcac gtgagcttca gctaccgcta cgaggccggc cgcgtgatcg
     3241 gcgacttcaa ggtggtgggc accggcttcc ccgaggacag cgtgatcttc accgacaaga
     3301 tcatccgcag caacgccacc gtggagcacc tgcaccccat gggcgataac gtgctggtgg
     3361 gcagcttcgc ccgcaccttc agcctgcgcg acggcggcta ctacagcttc gtggtggaca
     3421 gccacatgca cttcaagagc gccatccacc ccagcatcct gcagaacggg ggccccatgt
     3481 tcgccttccg ccgcgtggag gagctgcaca gcaacaccga gctgggcatc gtggagtacc
     3541 agcacgcctt caagaccccc attgccttcg ccagatcccg cgctcagtcg tccaattctg
     3601 ccgtggacgg caccgccgga cccggctcca ccggatctcg cgtcgacggc agtggagagg
     3661 gcagaggaag tctgctaaca tgcggtgacg tcgaggagaa tcctggccca atgaccgagt
     3721 acaagcccac ggtgcgcctc gccacccgcg acgacgtccc cagggccgta cgcaccctcg
     3781 ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgatccggac cgccacatcg
     3841 agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg
     3901 tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag agcgtcgaag
     3961 cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt tcccggctgg
     4021 ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt
     4081 tcctggccac cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc agcgccgtcg
     4141 tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg gagacctccg
     4201 cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg
     4261 tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc cggtgcctga gaattcgtcg
     4321 acaatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg
     4381 ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc
     4441 gtatggcttt cattttctcc tccttgtata aatcctggtt agttcttgcc acggcggaac
     4501 tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt
     4561 ccgtggtggg tacctttaag accaatgact tacaaggcag ctgtagatct tagccacttt
     4621 ttaaaagaaa aggggggact ggaagggcta attcactccc aacgaagata agatctgctt
     4681 tttgcttgta ctgggtctct ctggttagac cagatctgag cctgggagct ctctggctaa
     4741 ctagggaacc cactgcttaa gcctcaataa agcttgcctt gagtgcttca agtagtgtgt
     4801 gcccgtctgt tgtgtgactc tggtaactag agatccctca gaccctttta gtcagtgtgg
     4861 aaaatctcta gcagtagtag ttcatgtcat cttattattc agtatttata acttgcaaag
     4921 aaatgaatat cagagagtga gaggaacttg tttattgcag cttataatgg ttacaaataa
     4981 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     5041 ttgtccaaac tcatcaatgt atcttatcat gtctggctct agctatcccg cccctaactc
     5101 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     5161 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
     5221 gaggaggctt ttttggaggc ctagactttt gcagagacca aattcgtaat catgtcatag
     5281 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     5341 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     5401 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     5461 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     5521 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     5581 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     5641 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     5701 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     5761 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     5821 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     5881 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     5941 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     6001 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     6061 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     6121 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     6181 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     6241 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     6301 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     6361 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     6421 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     6481 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     6541 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     6601 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     6661 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     6721 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     6781 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     6841 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     6901 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     6961 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     7021 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     7081 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     7141 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     7201 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     7261 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     7321 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     7381 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc
     7441 attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg
     7501 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct
     7561 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc
     7621 gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat
     7681 atgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag gcgccattcg
     7741 ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc
     7801 cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc
     7861 cagtcacgac gttgtaaaac gacggccagt gccaagctg
//
