LOCUS       sequence-193267-        5192 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of eGFP in the presence of Cre.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5192)
  AUTHORS   Li Y, Kim J
  TITLE     Distinct roles of neuronal and microglial CB2 cannabinoid receptors 
            in the mouse hippocampus.
  JOURNAL   Neuroscience. 2017 Sep 6. pii: S0306-4522(17)30629-2. doi: 
            10.1016/j.neuroscience.2017.08.053.
  PUBMED    28888955
REFERENCE   2  (bases 1 to 5192)
  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: 
            "Neuroscience. 2017 Sep 6. pii: S0306-4522(17)30629-2. doi: 
            10.1016/j.neuroscience.2017.08.053."
COMMENT     Sequence Label: pAAV-CBh-DIO-EGFP
FEATURES             Location/Qualifiers
     source          1..5192
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     enhancer        145..430
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        432..709
                     /label=chicken beta-actin promoter
     intron          710..937
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     protein_bind    complement(967..1000)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(1051..1084)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(join(1112..1825,1826..1828,1829..1831))
                     /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             complement(join(1115..1825,1826..1828,1829..1831))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1157..1178)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1506..1525
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1765..1786
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      1828..1837
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    1860..1893
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    1944..1977
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(2047..2067)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   2455..2595
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2455..2584
                     /label=AAV2 ITR
     rep_origin      2670..3125
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2757..2776)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2967..2988
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3142..3161)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3261..3283
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3321..3339)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3407..3511
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3512..3580,3581..4372)
                     /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     complement(3730..3749)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4543..5131
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5032..5051
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtcgttac ataacttacg gtaaatggcc cgcctggctg
      181 accgcccaac gacccccgcc cattgacgtc aatagtaacg ccaataggga ctttccattg
      241 acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca
      301 tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattgtgc
      361 ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc
      421 tattaccatg gtcgaggtga gccccacgtt ctgcttcact ctccccatct cccccccctc
      481 cccaccccca attttgtatt tatttatttt ttaattattt tgtgcagcga tgggggcggg
      541 gggggggggg gggcgcgcgc caggcggggc ggggcggggc gaggggcggg gcggggcgag
      601 gcggagaggt gcggcggcag ccaatcagag cggcgcgctc cgaaagtttc cttttatggc
      661 gaggcggcgg cggcggcggc cctataaaaa gcgaagcgcg cggcgggcgg gagtcgctgc
      721 gcgctgcctt cgccccgtgc cccgctccgc cgccgcctcg cgccgcccgc cccggctctg
      781 actgaccgcg ttactcccac aggtgagcgg gcgggacggc ccttctcctc cgggctgtaa
      841 ttagctgagc aagaggtaag ggtttaaggg atggttggtt ggtggggtat taatgtttaa
      901 ttacctggag cacctgcctg aaatcacttt ttttcaggtt ggggatcctc tagagtcgac
      961 tccggaataa cttcgtatag gatactttat acgaagttat gcagaatggt agctggattg
     1021 tagctgctat tagcaatatg aaacctctta ataacttcgt atagcataca ttatacgaag
     1081 ttatggcgcg ccttatctag atttagccac cttacttgta cagctcgtcc atgccgagag
     1141 tgatcccggc ggcggtcacg aactccagca ggaccatgtg atcgcgcttc tcgttggggt
     1201 ctttgctcag ggcggactgg gtgctcaggt agtggttgtc gggcagcagc acggggccgt
     1261 cgccgatggg ggtgttctgc tggtagtggt cggcgagctg cacgctgccg tcctcgatgt
     1321 tgtggcggat cttgaagttc accttgatgc cgttcttctg cttgtcggcc atgatataga
     1381 cgttgtggct gttgtagttg tactccagct tgtgccccag gatgttgccg tcctccttga
     1441 agtcgatgcc cttcagctcg atgcggttca ccagggtgtc gccctcgaac ttcacctcgg
     1501 cgcgggtctt gtagttgccg tcgtccttga agaagatggt gcgctcctgg acgtagcctt
     1561 cgggcatggc ggacttgaag aagtcgtgct gcttcatgtg gtcggggtag cggctgaagc
     1621 actgcacgcc gtaggtcagg gtggtcacga gggtgggcca gggcacgggc agcttgccgg
     1681 tggtgcagat gaacttcagg gtcagcttgc cgtaggtggc atcgccctcg ccctcgccgg
     1741 acacgctgaa cttgtggccg tttacgtcgc cgtccagctc gaccaggatg ggcaccaccc
     1801 cggtgaacag ctcctcgccc ttgctcacca tggtggcggt ggcgaccggt agcgctagca
     1861 taacttcgta taaagtatcc tatacgaagt tatttgcctt aacccagaaa ttatcactgt
     1921 tattctttag aatggtgcaa agaataactt cgtataatgt atgctatacg aagttatgaa
     1981 ttccgctcga gataatcaac ctctggatta caaaatttgt gaaagattga ctggtattct
     2041 taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc
     2101 tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt tagttcttgc
     2161 cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg
     2221 cactgacaat tccgtggtgt ttatttgtga aatttgtgat gctattgctt tatttgtaac
     2281 catctagctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct
     2341 gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt cagggggaga
     2401 tgtgggaggt tttttaaagc ggccgcggta accacgtgcg gaccgagcgg ccgcaggaac
     2461 ccctagtgat ggagttggcc actccctctc tgcgcgctcg ctcgctcact gaggccgggc
     2521 gaccaaaggt cgcccgacgc ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc
     2581 gcagctgcct gcaggggcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     2641 cacaccgcat acgtcaaagc aaccatagta cgcgccctgt agcggcgcat taagcgcggc
     2701 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccttag cgcccgctcc
     2761 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     2821 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     2881 tgatttgggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     2941 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     3001 ctctatctcg ggctattctt ttgatttata agggattttg ccgatttcgg tctattggtt
     3061 aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgtttac
     3121 aattttatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagccccg
     3181 acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta
     3241 cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc
     3301 gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat
     3361 aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     3421 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     3481 aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct
     3541 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa
     3601 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa
     3661 cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt
     3721 taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg
     3781 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca
     3841 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa
     3901 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt
     3961 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc
     4021 cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa
     4081 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga
     4141 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc
     4201 tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga
     4261 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga
     4321 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga
     4381 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     4441 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     4501 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     4561 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     4621 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     4681 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     4741 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     4801 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     4861 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     4921 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     4981 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     5041 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     5101 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     5161 cctggccttt tgctggcctt ttgctcacat gt
//
