LOCUS       sequence-177275-        5934 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bicistronic AAV construct encoding GFP control and shRNA targeting 
            Ng.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5934)
  AUTHORS   Han KS, Cooke SF, Xu W
  TITLE     Experience-Dependent Equilibration of AMPAR-Mediated Synaptic 
            Transmission during the Critical Period.
  JOURNAL   Cell Rep. 2017 Jan 24;18(4):892-904. doi: 
            10.1016/j.celrep.2016.12.084.
  PUBMED    28122240
REFERENCE   2  (bases 1 to 5934)
  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: 
            "10.1016/j.celrep.2016.12"; journalName: "Cell Rep"; date: 
            "2017-01-24- 24"; volume: "18"; issue: "4"; pages: "892-904"
COMMENT     Sequence Label: shNg pA_RC3J1_CAGW
FEATURES             Location/Qualifiers
     source          1..5934
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     promoter        196..410
                     /label=H1 promoter
                     /note="human H1 RNA promoter"
     primer_bind     331..350
                     /label=H1
                     /note="Human H1 promoter, forward primer"
     promoter        743..1020
                     /label=chicken beta-actin promoter
     primer_bind     1300..1322
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1384..1403
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1455..1464
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1461..1463,1464..1466,1467..2180)
                     /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(1461..1463,1464..1466,1467..2177)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1506..1527)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1767..1786)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2114..2135
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     intron          2206..2338
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     2321..2340
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    2378..2966
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2431..2451)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2849..2860)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3009..3130
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3020..3039)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3074..3093
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   3292..3421
                     /label=AAV2 ITR
     rep_origin      complement(3658..4246)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3738..3757)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4417..5208,5209..5277))
                     /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     5040..5059
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5278..5382)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5408..5863)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5545..5566)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5757..5776
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct tgtagttaat gattaacccg ccatgctact tatctacgta gccatgctct
      181 aggaagatcg aattcgaacg ctgacgtcat caacccgctc caaggaatcg cgggcccagt
      241 gtcactaggc gggaacaccc agcgcgcgtg cgccctggca ggaagatggc tgtgagggac
      301 aggggagtgg cgccctgcaa tatttgcatg tcgctatgtg ttctgggaaa tcaccataaa
      361 cgtgaaatgt ctttggattt gggaatctta taagttctgt atgagaccac agatgcccgt
      421 gacaagactt ccctactgtt tcaagagaac agtagggaag tcttgtcact ttttggaaag
      481 cttatcgaag taccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg
      541 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat
      601 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc
      661 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta
      721 ttagtcatcg ctattaccat ggtcgaggtg agccccacgt tctgcttcac tctccccatc
      781 tcccccccct ccccaccccc aattttgtat ttatttattt tttaattatt ttgtgcagcg
      841 atgggggcgg gggggggggg ggggcgcgcg ccaggcgggg cggggcgggg cgaggggcgg
      901 ggcggggcga ggcggagagg tgcggcggca gccaatcaga gcggcgcgct ccgaaagttt
      961 ccttttatgg cgaggcggcg gcggcggcgg ccctataaaa agcgaagcgc gcggcgggcg
     1021 ggagtcgctg cgacgctgcc ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc
     1081 gccccggctc tgactgaccg cgttactccc acaggtgagc gggcgggacg gcccttctcc
     1141 tccgggctgt aattagcgct tggtttaatg acggcttgtt tcttttctgt ggctgcgtga
     1201 aagccttgag gggctccggg agggcccttt gtgcgggggg agcggctcgg ggctgtccgc
     1261 ggggggacgg ctgccttcgg gggggacggg gcagggcggg gttcggcttc tggcgtgtga
     1321 ccggcggctc tagagcctct gctaaccatg ttcatgcctt cttctttttc ctacagctcc
     1381 tgggcaacgt gctggttatt gtgctgtctc atcattttgg caaagaattg gatcgggatc
     1441 cccgggtacc ggtcgccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc
     1501 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg
     1561 gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc
     1621 tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc
     1681 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg
     1741 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga
     1801 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg
     1861 acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca
     1921 tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg
     1981 acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg
     2041 tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg
     2101 agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca
     2161 tggacgagct gtacaagtaa agcggccccg gactcgaggc cgcaggtaag tatcaaggtt
     2221 acaagacagg tttaaggaga ccaatagaaa ctgggcttgt cgagacagag aagactcttg
     2281 cgtttctgat aggcacctat tggtcttact gacatccact ttgcctttct ctccacaggt
     2341 gtcgagtgga gctcgcgact agatcaagct tatcgataat caacctctgg attacaaaat
     2401 ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc
     2461 tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt
     2521 gtataaatcc tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg
     2581 cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg
     2641 tcagctcctt tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc
     2701 cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt
     2761 gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc tgtgttgcca cctggattct
     2821 gcgcgggacg tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg
     2881 cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg
     2941 gatctccctt tgggccgcct ccccgcatcg ataccgtcga cccgggcggc cgcttcgagc
     3001 agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa
     3061 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa
     3121 taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg gggagatgtg
     3181 ggaggttttt taaagcaagt aaaacctcta caaatgtggt aaaatcgata aggatcttcc
     3241 tagagcatgg ctacgtagat aagtagcatg gcgggttaat cattaactac aaggaacccc
     3301 tagtgatgga gttggccact ccctctctgc gcgctcgctc gctcactgag gccgggcgac
     3361 caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc agtgagcgag cgagcgcgca
     3421 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     3481 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     3541 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     3601 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     3661 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     3721 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     3781 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     3841 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     3901 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     3961 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     4021 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     4081 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     4141 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     4201 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     4261 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     4321 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     4381 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     4441 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     4501 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     4561 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     4621 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     4681 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     4741 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     4801 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     4861 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     4921 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     4981 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     5041 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     5101 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     5161 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     5221 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     5281 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     5341 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     5401 gccacctaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
     5461 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
     5521 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
     5581 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
     5641 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
     5701 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg
     5761 aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta
     5821 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcccattc gccattcagg
     5881 ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccag
//
