LOCUS       sequence-341672-        4950 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  An AAV vector expressing shRNA vs rat PKCd and a nuclear EYFP 
            reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4950)
  AUTHORS   Venniro M, Russell TI, Ramsey LA, Richie CT, Lesscher HMB, 
            Giovanetti SM, Messing RO, Shaham Y
  TITLE     Abstinence-dependent dissociable central amygdala microcircuits 
            control drug craving.
  JOURNAL   Proc Natl Acad Sci U S A. 2020 Apr 7;117(14):8126-8134. doi: 
            10.1073/pnas.2001615117. Epub 2020 Mar 23.
  PUBMED    32205443
REFERENCE   2  (bases 1 to 4950)
  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.2001615117"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2020-04-7- 7"; volume: "117"; issue: "14"; pages: "8126-8134"
COMMENT     Sequence Label: pOTTC1534	pscAAV mU6 shRNA(PKCd) CMV-IE Nuc-EYFP
FEATURES             Location/Qualifiers
     source          1..4950
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(187..499)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     complement(248..268)
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     enhancer        594..897
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        898..1101
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1052..1072
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             join(1161..1163,1164..1166,1167..1877)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1206..1227)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1467..1486)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1814..1835
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1902..1922
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1926..1946
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(1984..2001)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1990..2101
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2213..2353
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2213..2342
                     /label=AAV2 ITR
     rep_origin      2428..2883
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2515..2534)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2725..2746
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2900..2919)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3019..3041
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3079..3097)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3165..3269
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3270..3338,3339..4130)
                     /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(3488..3507)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4301..4889
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4790..4809
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggctgc ggccgcacgc
      121 gtcgacctcg agaaaaaagg acaaaggccg cttcgaatct cttgaattcg aagcggcctt
      181 tgtccaaaca aggcttttct ccaagggata tttatagtct caaaacacac aattacttta
      241 cagttagggt gagtttcctt ttgtgctgtt ttttaaaata ataatttagt atttgtatct
      301 cttatagaaa tccaagccta tcatgtaaaa tgtagctagt attaaaaaga acagattatc
      361 tgtcttttat cgcacattaa gcctctatag ttactaggaa atattatatg caaattaacc
      421 ggggcagggg agtagccgag cttctcccac aagtctgtgc gagggggccg gcgcgggcct
      481 agagatggcg cgtcggatct ctagagcggg cccggtaacg cgtggatcta atatagttat
      541 taatagtaat caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca
      601 taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca
      661 ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg
      721 gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg
      781 ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc
      841 ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg
      901 atgcggtttt ggcagtacat caatgggcgt ggatagcggt ttgactcacg gggatttcca
      961 agtctccacc ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt
     1021 ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg
     1081 gaggtctata taagcagagc tggtttagtg aaccgtcaga tccgaagtta tcgctagcgc
     1141 taccggactc agatctcacc atggtgagca agggcgagga gctgttcacc ggggtggtgc
     1201 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg
     1261 gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc
     1321 tgcccgtgcc ctggcccacc ctcgtgacca ccttcggcta cggcctgcag tgcttcgccc
     1381 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg
     1441 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga
     1501 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg
     1561 acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca
     1621 tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg
     1681 acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg
     1741 tgctgctgcc cgacaaccac tacctgagct accagtccgc cctgagcaaa gaccccaacg
     1801 agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca
     1861 tggacgagct gtacaagtcc ggcctgcgca gccgcgccga cccaaaaaag aagcgcaagg
     1921 tagaccccaa gaagaagcgc aaggtagatc caaaaaagaa gagaaggtat gatagtctag
     1981 aagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt
     2041 ccttgaccct ggaaggtgcc actcccaccg tcctttccta ataaaatgag gaaattgcat
     2101 cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg
     2161 gggaggattg ggaagacaat agcaggcatg ctggggcgga ccgagcggcc gcaggaaccc
     2221 ctagtgatgg agttggccac tccctctctg cgcgctcgct cgctcactga ggccgggcga
     2281 ccaaaggtcg cccgacgccc gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc
     2341 agctgcctgc aggggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     2401 caccgcatac gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta agcgcggcgg
     2461 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccttagcg cccgctcctt
     2521 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     2581 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     2641 atttgggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     2701 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaact
     2761 ctatctcggg ctattctttt gatttataag ggattttgcc gatttcggtc tattggttaa
     2821 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgtttacaa
     2881 ttttatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagccccgac
     2941 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
     3001 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
     3061 aacgcgcgag acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa
     3121 taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     3181 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     3241 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     3301 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     3361 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     3421 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     3481 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     3541 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     3601 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     3661 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     3721 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     3781 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     3841 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     3901 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     3961 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     4021 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     4081 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     4141 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     4201 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     4261 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     4321 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     4381 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     4441 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     4501 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     4561 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     4621 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     4681 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     4741 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     4801 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     4861 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     4921 tggccttttg ctggcctttt gctcacatgt
//
