LOCUS       sequence-468746-        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 #1)
FEATURES             Location/Qualifiers
     source          1..6027
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(272..296)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        complement(293..496)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(322..342)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        497..800
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     repeat_region   876..1005
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(1067..1655)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1147..1166)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1826..2617,2618..2686))
                     /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     2449..2468
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2687..2791)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2859..2877
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2915..2937)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3037..3056
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(3073..3528)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3210..3231)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3422..3441
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   3603..3743
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3614..3743
                     /label=AAV2 ITR
     polyA_signal    3857..3991
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3894..3913)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3948..3967
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        complement(4070..4382)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     complement(4131..4151)
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     polyA_signal    4439..4915
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     4697..4716
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     CDS             complement(join(4968..5681,5682..5684,5685..5687))
                     /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(4971..5681,5682..5684,5685..5687))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(5013..5034)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     5362..5381
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5621..5642
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      5684..5693
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     intron          join(5793..6027,1..144)
                     /label=chimeric intron
                     /note="fusion between human cytomegalovirus intron A and 
                     human beta-globin intron 2"
ORIGIN
        1 gtgcaaagag gcatgataca ttgtatcatt attgccctga aagaaagaga ttagggaaag
       61 tattagaaat aagataaaca aaaaagtata ttaaaagaag aaagcatttt ttgtgggcct
      121 atagactcta taggcggtac ttacgtcact cttggcacgg ggaatccgcg ttccaatgca
      181 ccgttcccgg ccgggattcg aatccgcgga ggctggatcg gtcccggtgt cttctatgga
      241 ggtcaaaaca gcgtggatgg cgtctccagg cgatctgacg gttcactaaa cgagctctgc
      301 ttatatagac ctcccaccgt acacgcctac cgcccatttg cgtcaatggg gcggagttgt
      361 tacgacattt tggaaagtcc cgttgatttt ggtgccaaaa caaactccca ttgacgtcaa
      421 tggggtggag acttggaaat ccccgtgagt caaaccgcta tccacgccca ttgatgtact
      481 gccaaaaccg catcaccatg gtaatagcga tgactaatac gtagatgtac tgccaagtag
      541 gaaagtccca taaggtcatg tactgggcat aatgccaggc gggccattta ccgtcattga
      601 cgtcaatagg gggcgtactt ggcatatgat acacttgatg tactgccaag tgggcagttt
      661 accgtaaata ctccacccat tgacgtcaat ggaaagtccc tattggcgtt actatgggaa
      721 catacgtcat tattgacgtc aatgggcggg ggtcgttggg cggtcagcca ggcgggccat
      781 ttaccgtaag ttatgtaacg cggaactcca tatatgggct atgaactaat gaccccgtaa
      841 ttgattacta ttaataacta gacgcgtgcg gccgcaggaa cccctagtga tggagttggc
      901 cactccctct ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg
      961 cccgggcggc ctcagtgagc gagcgagcgc gcagctgcct gcaggacatg tgagcaaaag
     1021 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     1081 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     1141 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     1201 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     1261 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     1321 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     1381 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     1441 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     1501 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     1561 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     1621 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     1681 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     1741 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     1801 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     1861 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     1921 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     1981 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     2041 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     2101 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     2161 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     2221 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     2281 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     2341 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     2401 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     2461 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     2521 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     2581 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     2641 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     2701 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     2761 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
     2821 tctaagaaac cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct
     2881 ttcgtctcgc gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga
     2941 cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag
     3001 cgggtgttgg cgggtgtcgg ggctggctta actatgcggc atcagagcag attgtactga
     3061 gagtgcacca taaaattgta aacgttaata ttttgttaaa attcgcgtta aatttttgtt
     3121 aaatcagctc attttttaac caatagaccg aaatcggcaa aatcccttat aaatcaaaag
     3181 aatagcccga gatagagttg agtgttgttc cagtttggaa caagagtcca ctattaaaga
     3241 acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg
     3301 aaccatcacc caaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc
     3361 ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg
     3421 aagggaagaa agcgaaagga gcgggcgcta aggcgctggc aagtgtagcg gtcacgctgc
     3481 gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtac tatggttgct
     3541 ttgacgtatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc gcatcaggcg
     3601 cccctgcagg cagctgcgcg ctcgctcgct cactgaggcc gcccgggcaa agcccgggcg
     3661 tcgggcgacc tttggtcgcc cggcctcagt gagcgagcga gcgcgcagag agggagtggc
     3721 caactccatc actaggggtt cctgcggccg ctcggtccgc acgtggttac cggtaccgcc
     3781 gcggccgcaa gaattaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa
     3841 tgcaattgtt gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     3901 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     3961 actcatcaat gtatcttatc atgtctggat ctacgtatta tctagaaaaa taaatcggag
     4021 ctgtattctc ccgatgacag gaagtcggga gaatacagct ccgatttcaa acaaggcttt
     4081 tctccaaggg atatttatag tctcaaaaca cacaattact ttacagttag ggtgagtttc
     4141 cttttgtgct gttttttaaa ataataattt agtatttgta tctcttatag aaatccaagc
     4201 ctatcatgta aaatgtagct agtattaaaa agaacagatt atctgtcttt tatcgcacat
     4261 taagcctcta tagttactag gaaatattat atgcaaatta accggggcag gggagtagcc
     4321 gagcttctcc cacaagtctg tgcgaggggg ccggcgcggg cctagagatg gcggcgtcgg
     4381 ataagcttgg cctagatggc cttggcgcgc cattgggccc ggttacctac aaaatcagaa
     4441 ggacagggaa gggagcagtg gttcacgcct gtaatcccag caatttggga ggccaaggtg
     4501 ggtagatcac ctgagattag gagttggaga ccagcctggc caatatggtg aaaccccgtc
     4561 tctaccaaaa aaacaaaaat tagctgagcc tggtcatgca tgcctggaat cccaacaact
     4621 cgggaggctg aggcaggaga atcgcttgaa cccaggaggc ggagattgca gtgagccaag
     4681 attgtgccac tgcactccag cttggttccc aatagacccc gcaggcccta caggttgtct
     4741 tcccaacttg ccccttgctc cataccaccc ccctccaccc cataatatta tagaaggaca
     4801 cctagtcaga caaaatgatg caacttaatt ttattaggac aaggctggtg ggcactggag
     4861 tggcaacttc cagggccagg agaggcactg gggaggggtc acagggatgc cacccgtaga
     4921 tctctcgagc agcgctgctc gaggcaagct tctgcaggtc ggccgcttta cttgtacagc
     4981 tcgtccatgc cgagagtgat cccggcggcg gtcacgaact ccagcaggac catgtgatcg
     5041 cgcttctcgt tggggtcttt gctcagggcg gactgggtgc tcaggtagtg gttgtcgggc
     5101 agcagcacgg ggccgtcgcc gatgggggtg ttctgctggt agtggtcggc gagctgcacg
     5161 ctgccgtcct cgatgttgtg gcggatcttg aagttcacct tgatgccgtt cttctgcttg
     5221 tcggccatga tatagacgtt gtggctgttg tagttgtact ccagcttgtg ccccaggatg
     5281 ttgccgtcct ccttgaagtc gatgcccttc agctcgatgc ggttcaccag ggtgtcgccc
     5341 tcgaacttca cctcggcgcg ggtcttgtag ttgccgtcgt ccttgaagaa gatggtgcgc
     5401 tcctggacgt agccttcggg catggcggac ttgaagaagt cgtgctgctt catgtggtcg
     5461 gggtagcggc tgaagcactg cacgccgtag gtcagggtgg tcacgagggt gggccagggc
     5521 acgggcagct tgccggtggt gcagatgaac ttcagggtca gcttgccgta ggtggcatcg
     5581 ccctcgccct cgccggacac gctgaacttg tggccgttta cgtcgccgtc cagctcgacc
     5641 aggatgggca ccaccccggt gaacagctcc tcgcccttgc tcaccatggt ggcgaccggt
     5701 ggatccccga ctgcagaatt caatcgatgt tcgaatccca attctttgcc aaagtgatgg
     5761 gccagcacac agaccagcac gttgcccagg agctgtggga ggaagataag aggtatgaac
     5821 atgattagca aaagggccta gcttggactc agaataatcc agccttatcc caaccataaa
     5881 ataaaagcag aatggtagct ggattgtagc tgctattagc aatatgaaac ctcttacatc
     5941 agttacaatt tatatgcaga aatatttata tgcagaaata ttgctattgc cttaacccag
     6001 aaattatcac tgttattctt tagaatg
//
