LOCUS       sequence-229299-        6913 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of hM4D(Gi) using the CD68 promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6913)
  AUTHORS   Grace PM, Strand KA, Galer EL, Urban DJ, Wang X, Baratta MV, 
            Fabisiak TJ, Anderson ND, Cheng K, Greene LI, Berkelhammer D, Zhang 
            Y, Ellis AL, Yin HH, Campeau S, Rice KC, Roth BL, Maier SF, Watkins 
            LR
  TITLE     Morphine paradoxically prolongs neuropathic pain in rats by 
            amplifying spinal NLRP3 inflammasome activation.
  JOURNAL   Proc Natl Acad Sci U S A. 2016 Jun 14;113(24):E3441-50. doi: 
            10.1073/pnas.1602070113. Epub 2016 May 31.
  PUBMED    27247388
REFERENCE   2  (bases 1 to 6913)
  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"; doi: 
            "10.1073/pnas.1602070113"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2016-06-14- 14"; volume: "113"; issue: "24"; pages: 
            "E3441-50"
COMMENT     Sequence Label: pAAV CD68-hM4D(Gi)-mCherry
FEATURES             Location/Qualifiers
     source          1..6913
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(26..47)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     238..257
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   419..559
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   430..559
                     /label=AAV2 ITR
     polyA_signal    599..1075
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     857..876
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     misc_feature    1107..1695
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             1213..1224
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1622..1642
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1720..2430)
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1998..2017)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2267..2285
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     regulatory      2427..2436
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      3885..3894
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     repeat_region   4605..4734
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(4796..5384)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4876..4895)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5555..6346,6347..6415))
                     /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     6178..6197
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6416..6520)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6588..6606
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6644..6666)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6766..6785
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(join(6802..6913,1..344))
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
ORIGIN
        1 gcccgagata gagttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt
       61 ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc
      121 atcacccaaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa
      181 agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg
      241 gaagaaagcg aaaggagcgg gcgctaaggc gctggcaagt gtagcggtca cgctgcgcgt
      301 aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtactatg gttgctttga
      361 cgtatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat caggcgcccc
      421 tgcaggcagc tgcgcgctcg ctcgctcact gaggccgccc gggcaaagcc cgggcgtcgg
      481 gcgacctttg gtcgcccggc ctcagtgagc gagcgagcgc gcagagaggg agtggccaac
      541 tccatcacta ggggttcctg cggccgctcg gtccgcacgt ggttacctac aaaatcagaa
      601 ggacagggaa gggagcagtg gttcacgcct gtaatcccag caatttggga ggccaaggtg
      661 ggtagatcac ctgagattag gagttggaga ccagcctggc caatatggtg aaaccccgtc
      721 tctaccaaaa aaacaaaaat tagctgagcc tggtcatgca tgcctggaat cccaacaact
      781 cgggaggctg aggcaggaga atcgcttgaa cccaggaggc ggagattgca gtgagccaag
      841 attgtgccac tgcactccag cttggttccc aatagacccc gcaggcccta caggttgtct
      901 tcccaacttg ccccttgctc cataccaccc ccctccaccc cataatatta tagaaggaca
      961 cctagtcaga caaaatgatg caacttaatt ttattaggac aaggctggtg ggcactggag
     1021 tggcaacttc cagggccagg agaggcactg gggaggggtc acagggatgc cacccgtaga
     1081 tctctcgagc agcgctcggt atcgatgcgg ggaggcggcc caaagggaga tccgactcgt
     1141 ctgagggcga aggcgaagac gcggaagagg ccgcagagcc ggcagcaggc cgcgggaagg
     1201 aaggtccgct ggattgaggg ccgaagggac gtagcagaag gacgtcccgc gcagaatcca
     1261 ggtggcaaca taggcgagca gccaaggaaa ggacgatgat ttccccgaca acaccacgga
     1321 attgtcagtg cccaacagcc gagcccctgt ccagcagcgg gcaaggcagg cggcgatgag
     1381 ttccgccgtg gcaataggga gggggaaagc gaaagtcccg gaaaggagct gacaggtggt
     1441 ggcaatgccc caaccagtgg gggttgcgtc agcaaacaca gtgcacacca cgccacgttg
     1501 cctgacaacg ggccacaact cctcataaag agacagcaac caggatttat acaaggagga
     1561 gaaaatgaaa gccatacggg aagcaatagc atgatacaaa ggcattaaag cagcgtatcc
     1621 acatagcgta aaaggagcaa catagttaag aataccagtc aatctttcac aaattttgta
     1681 atccagaggt tgattatcga taagcttgat atcgaattct tacttgtaca gctcgtccat
     1741 gccgccggtg gagtggcggc cctcggcgcg ttcgtactgt tccacgatgg tgtagtcctc
     1801 gttgtgggag gtgatgtcca acttgatgtt gacgttgtag gcgccgggca gctgcacggg
     1861 cttcttggcc ttgtaggtgg tcttgacctc agcgtcgtag tggccgccgt ccttcagctt
     1921 cagcctctgc ttgatctcgc ccttcagggc gccgtcctcg gggtacatcc gctcggagga
     1981 ggcctcccag cccatggtct tcttctgcat tacggggccg tcggagggga agttggtgcc
     2041 gcgcagcttc accttgtaga tgaactcgcc gtcctgcagg gaggagtcct gggtcacggt
     2101 caccacgccg ccgtcctcga agttcatcac gcgctcccac ttgaagccct cggggaagga
     2161 cagcttcaag tagtcgggga tgtcggcggg gtgcttcacg taggccttgg agccgtacat
     2221 gaactgaggg gacaggatgt cccaggcgaa gggcaggggg ccacccttgg tcaccttcag
     2281 cttggcggtc tgggtgccct cgtaggggcg gccctcgccc tcgccctcga tctcgaactc
     2341 gtggccgttc acggagccct ccatgtgcac cttgaagcgc atgaactcct tgatgatggc
     2401 catgttatcc tcctcgccct tgctcaccat ggtggcgacc ggtggatccc gcctggcagt
     2461 gccgatgttc cgatactggc acagcagcag gtgccggaag gtctttttaa aggtggcgtt
     2521 gcacagagca tagcaggcag ggttgatggt gctgttgacg tagcagagcc agtagccaat
     2581 ggaccacacc gtgtcaggga tgcagctctg gcagaaggtg ttcaccagga ccatgacgtt
     2641 gtagggcgtc caggtgagga tgaaggccag cagaatggca aagatcgttc gtgtcacttt
     2701 gcgctcccgg gccgccatct gccgcttctt gcgcacctgg ttgcgagcga tgctggcgaa
     2761 cttgcgggcc acgttggccg cagggcgcat gccagccggc gtggcaggca caatctcaat
     2821 ggctgtcaca cactcattgc ctgtctgctt cgtcacaatc tggatcttgg accatctgga
     2881 ggctgggttg agggcccgcg gctgcagggg aggggcgggc atggcgggcg tggtggcctc
     2941 tgtggtggac agctctgtgg ctgggcgttc cttggtgttc tgggtggcac tgcctgagct
     3001 ggactcattg gaagtgtcct tatcagccac ggggcgcggt ggcggtggca gcgctggcgg
     3061 gggggcctcc tccagcttgc cattgcgcag ctcctcccgg gcggcctccc cgggcggggg
     3121 cttcttgacg ctctgcttca ttagtgggct cttgaggaag gccagcgtct tggctttctt
     3181 ctccttcggg ccctcgggcc ggtgcttgtg gactcggctg cgactggcca gggagatgtg
     3241 gatgtacagc accgtcatga tgaccacagg caggtagaag ccagcaatgg ctgtgccaaa
     3301 ggtcactgct gggttggaca ggaactggat gaagcactgg ttgtcgggca ccgtccgctt
     3361 acccaccaca aactgccaga acaagatggc aggcgcccag agcacgaagg acagtaccca
     3421 ggcagcagca atcatgaggc ctgccatctt ggtggtgcgc cgggcagggt aggtgagagg
     3481 cttggtgacg cagaagtagc ggtcaaagct gatgatgaga aggttcatga cggaggcgtt
     3541 gctcaccacg cagtccaggg ccagccacag gtcgcagacc acggcgccca ggggccagta
     3601 gcccttgatg atgtacacgg tgtagaggtt catggagaag gcgcctatga tgagatcagc
     3661 acacgccagg ctgaagagga agtagttgtt gactgtctgc agctgcctgt tgaccttgat
     3721 ggacagcatc accaggatgt tgcccacgac agtcaccagg ctcagggagc ctgtcactgt
     3781 ggcaatgaag accatttcca ccgtctcata gcgattgtgg gatgatgacg tgaccaggcg
     3841 cacggactga ttgcccgagc tgccattgac aggtgtgaag ttggccatgg tggcgtcgac
     3901 gatatggctg aaccgcctca cccaaagtcc agctaggctg tctgcaccag ttgctcaacc
     3961 ctgctccctc agccctctct tggaaaggag gaaatgaaag tctcacttcc tgtactggag
     4021 atgatcccac ccacagcctg actcaggggc ctcagctcta gccacctctc tcttagtcat
     4081 tagtttttgt aattaagtta ggggactgtt ggccctgctt cgtcagggca aagtcaacag
     4141 tgtcacatgg gtgcaaacgg agtcctaggg cagttgatcc acagggatcc atgtgtcaaa
     4201 ctcagcgaaa cttgcttggc agttgtggca agtagctatg ggaacagctg aggtacaggc
     4261 ccctcccttg gtggggaaat cggtttccac tcctttcccc tggacctctg ggctgacccc
     4321 tgtactactt cctcttggct gaggcatgtg aggggcttgt gaccatccac acccagccct
     4381 ttacagaggg gcagcactcc agtcagcatt gtcattcaca gagccgagaa catggctttc
     4441 cagcgtctga gaactcagat ggtgacaaga aagcaacagc ccaaaggtaa ccatctgctg
     4501 gggaaggggg gagcagagat gcacccagta gcaggtggga gagggaaggg gaggtaagaa
     4561 atgagaagcc caaacaggca gtttgatatc acgcgtgcgg ccgcaggaac ccctagtgat
     4621 ggagttggcc actccctctc tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt
     4681 cgcccgacgc ccgggcggcc tcagtgagcg agcgagcgcg cagctgcctg caggacatgt
     4741 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     4801 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     4861 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     4921 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     4981 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     5041 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     5101 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     5161 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     5221 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     5281 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     5341 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     5401 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     5461 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     5521 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     5581 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     5641 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     5701 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     5761 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     5821 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     5881 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     5941 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     6001 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     6061 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     6121 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     6181 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     6241 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     6301 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     6361 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     6421 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     6481 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     6541 cacctgacgt ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca
     6601 cgaggccctt tcgtctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc
     6661 tcccggagac ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg
     6721 gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa ctatgcggca tcagagcaga
     6781 ttgtactgag agtgcaccat aaaattgtaa acgttaatat tttgttaaaa ttcgcgttaa
     6841 atttttgtta aatcagctca ttttttaacc aatagaccga aatcggcaaa atcccttata
     6901 aatcaaaaga ata
//
