LOCUS       sequence-353877-        5783 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express human cannabinoid 2 receptor (GFP tagged) in mammalian 
            cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5783)
  AUTHORS   Tomasevic N, Emser FS, Muratspahic E, Gattringer J, Hasinger S, 
            Hellinger R, Keov P, Felkl M, Gertsch J, Becker CFW, Gruber CW
  TITLE     Discovery and development of macrocyclic peptide modulators of the 
            cannabinoid 2 receptor.
  JOURNAL   J Biol Chem. 2024 Jun;300(6):107330. doi: 10.1016/j.jbc.2024.107330.
            Epub 2024 Apr 26.
  PUBMED    38679329
REFERENCE   2  (bases 1 to 5783)
  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: "J Biol 
            Chem."; date: "2024-06"; volume: "300(6):107330. doi"; pages: " 
            10.1016/j.jbc.2024.107330. Epub 2024 Apr 26"
COMMENT     Sequence Label: pEGFP-hCB2R
FEATURES             Location/Qualifiers
     source          1..5783
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1723..1732
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1729..1731,1732..1734,1735..2448)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1729..1731,1732..1734,1735..2445)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1774..1795)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2035..2054)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2382..2403
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2571..2692
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2608..2627)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2662..2681
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2699..3154)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2836..2857)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3048..3067
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3181..3285
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3287..3644
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3287..3478
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3310..3330)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3495..3630
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3679..4473
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3679..4473
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3733..3752)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4343..4362
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4661..4680)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4705..4752
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5081..5669
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5570..5589
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgagct caagcttatg gaggaatgct gggtgacaga gatagccaat
      661 ggctccaagg atggcttgga ttccaaccct atgaaggatt acatgatcct gagtggtccc
      721 cagaagacag ctgttgctgt gttgtgcact cttctgggcc tgctaagtgc cctggagaac
      781 gtggctgtgc tctatctgat cctgtcctcc caccaactcc gccggaagcc ctcatacctg
      841 ttcattggca gcttggctgg ggctgacttc ctggccagtg tggtctttgc atgcagcttt
      901 gtgaatttcc atgttttcca tggtgtggat tccaaggctg tcttcctgct gaagattggc
      961 agcgtgacta tgaccttcac agcctctgtg ggtagcctcc tgctgaccgc cattgaccga
     1021 tacctctgcc tgcgctatcc accttcctac aaagctctgc tcacccgtgg aagggcactg
     1081 gtgaccctgg gtatcatgtg ggtcctctca gcactagtct cctacctgcc cctcatggga
     1141 tggacttgct gtcccaggcc ctgctctgag cttttcccac tgatccccaa tgactacctg
     1201 ctgagctggc tcctgttcat cgccttcctc ttttccggaa tcatctacac ctatgggcat
     1261 gttctctgga aggcccatca gcatgtagcc agcttgtctg gccaccagga caggcaggtg
     1321 ccaggaatgg cccgaatgag gctggatgtg aggttggcca agaccctagg gctagtgctg
     1381 gctgtgctcc tcatctgttg gttcccagtg ctggccctca tggcccacag cctggccact
     1441 acgctcagtg accaggtcaa gaaggccttt gccttctgct ccatgctgtg cctcatcaac
     1501 tccatggtca accctgtcat ctatgctcta cggagtggag agatccgctc ctctgcccat
     1561 cactgcctgg ctcactggaa gaagtgtgtg aggggccttg ggtcagaggc aaaagaagaa
     1621 gccccaagat cctcagtcac ggagacagag gctgatggga aaatcactcc gtggccagat
     1681 tccagagatc tagacctctc tgattgcaag gatccaccgg tcgccaccat ggtgagcaag
     1741 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
     1801 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
     1861 ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
     1921 ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc
     1981 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac
     2041 ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc
     2101 gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac
     2161 aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg
     2221 aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag
     2281 cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc
     2341 cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc
     2401 gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagtaaag cggccgcgac
     2461 tctagatcat aatcagccat accacatttg tagaggtttt acttgcttta aaaaacctcc
     2521 cacacctccc cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt aacttgttta
     2581 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     2641 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct taaggcgtaa
     2701 attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt
     2761 tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata
     2821 gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac
     2881 gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa
     2941 tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc
     3001 cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg
     3061 aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca
     3121 cccgccgcgc ttaatgcgcc gctacagggc gcgtcaggtg gcacttttcg gggaaatgtg
     3181 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     3241 caataaccct gataaatgct tcaataatat tgaaaaagga agagtcctga ggcggaaaga
     3301 accagctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
     3361 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     3421 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     3481 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     3541 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc
     3601 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaagatcga tcaagagaca
     3661 ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct
     3721 tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc
     3781 gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc
     3841 ggtgccctga atgaactgca agacgaggca gcgcggctat cgtggctggc cacgacgggc
     3901 gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg
     3961 ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc
     4021 atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac
     4081 caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat
     4141 caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc
     4201 aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg
     4261 aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg
     4321 gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc
     4381 gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc
     4441 gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg
     4501 accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa
     4561 ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc
     4621 tcatgctgga gttcttcgcc caccctaggg ggaggctaac tgaaacacgg aaggagacaa
     4681 taccggaagg aacccgcgct atgacggcaa taaaaagaca gaataaaacg cacggtgttg
     4741 ggtcgtttgt tcataaacgc ggggttcggt cccagggctg gcactctgtc gataccccac
     4801 cgagacccca ttggggccaa tacgcccgcg tttcttcctt ttccccaccc caccccccaa
     4861 gttcgggtga aggcccaggg ctcgcagcca acgtcggggc ggcaggccct gccatagcct
     4921 caggttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     4981 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     5041 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     5101 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     5161 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     5221 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     5281 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     5341 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     5401 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     5461 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     5521 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     5581 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     5641 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     5701 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     5761 ataaccgtat taccgccatg cat
//
