LOCUS       sequence-468743-        6027 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To express GFP from a CMV promoter and to express an shRNA targeting
            Rat Nurr1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6027)
  TITLE     Ploski Lab Plasmids 2025
REFERENCE   2  (bases 1 to 6027)
  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: pAAV-CMV-GFP-pA U6 shRNA (Rat Nurr1 #5)
FEATURES             Location/Qualifiers
     source          1..6027
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      337..346
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(343..345,346..348,349..1062)
                     /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(343..345,346..348,349..1059)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(388..409)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(649..668)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     996..1017
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1115..1591
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(1314..1333)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     promoter        1648..1960
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     1879..1899
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     polyA_signal    2039..2173
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2063..2082)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2117..2136
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   2287..2427
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2287..2416
                     /label=AAV2 ITR
     rep_origin      2502..2957
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2589..2608)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2799..2820
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2974..2993)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3093..3115
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3153..3171)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3239..3343
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3344..3412,3413..4204)
                     /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(3562..3581)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4375..4963
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4864..4883
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   5025..5154
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     enhancer        5230..5533
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5534..5737
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5688..5708
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     5734..5758
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          join(5886..6027,1..237)
                     /label=chimeric intron
                     /note="fusion between human cytomegalovirus intron A and 
                     human beta-globin intron 2"
ORIGIN
        1 accattctaa agaataacag tgataatttc tgggttaagg caatagcaat atttctgcat
       61 ataaatattt ctgcatataa attgtaactg atgtaagagg tttcatattg ctaatagcag
      121 ctacaatcca gctaccattc tgcttttatt ttatggttgg gataaggctg gattattctg
      181 agtccaagct aggccctttt gctaatcatg ttcatacctc ttatcttcct cccacagctc
      241 ctgggcaacg tgctggtctg tgtgctggcc catcactttg gcaaagaatt gggattcgaa
      301 catcgattga attctgcagt cggggatcca ccggtcgcca ccatggtgag caagggcgag
      361 gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac
      421 aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag
      481 ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc
      541 tacggcgtgc agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag
      601 tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac
      661 tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg
      721 aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac
      781 aacagccaca acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc
      841 aagatccgcc acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac
      901 acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc
      961 gccctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc
     1021 gccgccggga tcactctcgg catggacgag ctgtacaagt aaagcggccg acctgcagaa
     1081 gcttgcctcg agcagcgctg ctcgagagat ctacgggtgg catccctgtg acccctcccc
     1141 agtgcctctc ctggccctgg aagttgccac tccagtgccc accagccttg tcctaataaa
     1201 attaagttgc atcattttgt ctgactaggt gtccttctat aatattatgg ggtggagggg
     1261 ggtggtatgg agcaaggggc aagttgggaa gacaacctgt agggcctgcg gggtctattg
     1321 ggaaccaagc tggagtgcag tggcacaatc ttggctcact gcaatctccg cctcctgggt
     1381 tcaagcgatt ctcctgcctc agcctcccga gttgttggga ttccaggcat gcatgaccag
     1441 gctcagctaa tttttgtttt tttggtagag acggggtttc accatattgg ccaggctggt
     1501 ctccaactcc taatctcagg tgatctaccc accttggcct cccaaattgc tgggattaca
     1561 ggcgtgaacc actgctccct tccctgtcct tctgattttg taggtaaccg ggcccaatgg
     1621 cgcgccaagg ccatctaggc caagcttatc cgacgccgcc atctctaggc ccgcgccggc
     1681 cccctcgcac agacttgtgg gagaagctcg gctactcccc tgccccggtt aatttgcata
     1741 taatatttcc tagtaactat agaggcttaa tgtgcgataa aagacagata atctgttctt
     1801 tttaatacta gctacatttt acatgatagg cttggatttc tataagagat acaaatacta
     1861 aattattatt ttaaaaaaca gcacaaaagg aaactcaccc taactgtaaa gtaattgtgt
     1921 gttttgagac tataaatatc ccttggagaa aagccttgtt tgaagagtgg taactgtagc
     1981 tctgacttcc tgtcatcaga gctacagtta ccactcttat ttttctagat aatacgtaga
     2041 tccagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa
     2101 aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg
     2161 caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc agggggaggt
     2221 gtgggaggtt ttttaattct tgcggccgcg gcggtaccgg taaccacgtg cggaccgagc
     2281 ggccgcagga acccctagtg atggagttgg ccactccctc tctgcgcgct cgctcgctca
     2341 ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga
     2401 gcgagcgagc gcgcagctgc ctgcaggggc gcctgatgcg gtattttctc cttacgcatc
     2461 tgtgcggtat ttcacaccgc atacgtcaaa gcaaccatag tacgcgccct gtagcggcgc
     2521 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgcctt
     2581 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     2641 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     2701 ccccaaaaaa cttgatttgg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     2761 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     2821 aacaacactc aactctatct cgggctattc ttttgattta taagggattt tgccgatttc
     2881 ggtctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat
     2941 attaacgttt acaattttat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt
     3001 aagccagccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc
     3061 ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc
     3121 accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat ttttataggt
     3181 taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg
     3241 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     3301 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     3361 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     3421 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     3481 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     3541 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     3601 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     3661 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     3721 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     3781 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     3841 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
     3901 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     3961 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     4021 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     4081 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     4141 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     4201 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     4261 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     4321 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     4381 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     4441 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     4501 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     4561 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     4621 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     4681 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     4741 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     4801 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     4861 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     4921 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     4981 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgtcctgca ggcagctgcg
     5041 cgctcgctcg ctcactgagg ccgcccgggc gtcgggcgac ctttggtcgc ccggcctcag
     5101 tgagcgagcg agcgcgcaga gagggagtgg ccaactccat cactaggggt tcctgcggcc
     5161 gcacgcgtct agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat
     5221 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
     5281 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
     5341 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
     5401 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
     5461 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
     5521 cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga
     5581 ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca
     5641 aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg
     5701 taggcgtgta cggtgggagg tctatataag cagagctcgt ttagtgaacc gtcagatcgc
     5761 ctggagacgc catccacgct gttttgacct ccatagaaga caccgggacc gatccagcct
     5821 ccgcggattc gaatcccggc cgggaacggt gcattggaac gcggattccc cgtgccaaga
     5881 gtgacgtaag taccgcctat agagtctata ggcccacaaa aaatgctttc ttcttttaat
     5941 atactttttt gtttatctta tttctaatac tttccctaat ctctttcttt cagggcaata
     6001 atgatacaat gtatcatgcc tctttgc
//
