LOCUS       sequence-444906-        6422 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  shRNAmir backbone for RNA interference, with YFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6422)
  AUTHORS   Kim J, de Haro M, Al-Ramahi I, Garaicoechea LL, Jeong HH, Sonn JY, 
            Tadros B, Liu Z, Botas J, Zoghbi HY
  TITLE     Evolutionarily conserved regulators of tau identify targets for new 
            therapies.
  JOURNAL   Neuron. 2023 Mar 15;111(6):824-838.e7. doi: 
            10.1016/j.neuron.2022.12.012. Epub 2023 Jan 6.
  PUBMED    36610398
REFERENCE   2  (bases 1 to 6422)
  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.neuron.2022.12.012"; journalName: "Neuron"; date: 
            "2023-03-15- 15"; volume: "111"; issue: "6"; pages: "824-838.e7"
COMMENT     Sequence Label: Luciferase shRNA
FEATURES             Location/Qualifiers
     source          1..6422
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(55..846,847..915))
                     /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     678..697
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(916..1020)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1088..1106
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1144..1166)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     1266..1285
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1499..1518
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1808..1826
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1808..1825
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     repeat_region   1847..1976
                     /label=AAV2 ITR
     promoter        2282..2559
                     /label=chicken beta-actin promoter
     primer_bind     2838..2860
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     2922..2941
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      2981..2990
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2987..2989,2990..2992,2993..3706)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3032..3053)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3293..3312)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3640..3661
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     ncRNA           3892..3906
                     /label=miR-30a loop
                     /note="loop from the the 71-nt precursor of the human 
                     miR-30a microRNA (Zeng et al., 2002)"
                     /ncRNA_class="miRNA"
     misc_feature    4088..4676
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4141..4161)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4559..4570)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4703..4720)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4709..4933
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        complement(5143..5472)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5157..5292
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5457..5477
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(5718..6306)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5798..5817)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
       61 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
      121 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
      181 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
      241 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
      301 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
      361 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
      421 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
      481 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
      541 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
      601 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
      661 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
      721 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
      781 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
      841 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
      901 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
      961 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     1021 cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca tgacattaac
     1081 ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg atgacggtga
     1141 aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg
     1201 gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa
     1261 ctatgcggca tcagagcaga ttgtactgag agtgcaccat tcgacgctct cccttatgcg
     1321 actcctgcat taggaagcag cccagtagta ggttgaggcc gttgagcacc gccgccgcaa
     1381 ggaatggtgc atgcaaggag atggcgccca acagtccccc ggccacgggg cctgccacca
     1441 tacccacgcc gaaacaagcg ctcatgagcc cgaagtggcg agcccgatct tccccatcgg
     1501 tgatgtcggc gatataggcg ccagcaaccg cacctgtggc gccggtgatg ccggccacga
     1561 tgcgtccggc gtagaggatc tggctagcga tgaccctgct gattggttcg ctgaccattt
     1621 ccgggtgcgg gacggcgtta ccagaaactc agaaggttcg tccaaccaaa ccgactctga
     1681 cggcagttta cgagagagat gatagggtct gcttcagtaa gccagatgct acacaattag
     1741 gcttgtacat attgtcgtta gaacgcggct acaattaata cataacctta tgtatcatac
     1801 acatacgatt taggtgacac tatagaatac acggaattaa ttctagctgc gcgctcgctc
     1861 gctcactgag gccgcccggg caaagcccgg gcgtcgggcg acctttggtc gcccggcctc
     1921 agtgagcgag cgagcgcgca gagagggagt ggccaactcc atcactaggg gttccttgta
     1981 gttaatgatt aacccgccat gctacttatc tacgtagcca tgctctaggt accattgacg
     2041 tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg
     2101 gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt
     2161 acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg
     2221 accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg
     2281 gtcgaggtga gccccacgtt ctgcttcact ctccccatct cccccccctc cccaccccca
     2341 attttgtatt tatttatttt ttaattattt tgtgcagcga tgggggcggg gggggggggg
     2401 gggcgcgcgc cggggggggg ggggsggggs grggggsggg gsggggsgrg gcggagaggt
     2461 gcggcggcag ccaatcagag cggcgcgctc cgaaagtttc cttttatggc gaggcggcgg
     2521 cggcggcggc cctataaaaa gcgaagcgcg cggcgggcgg gagtcgctgc gcgctgcctt
     2581 cgccccgtgc cccgctccgc cgccgcctcg cgccgcccgc cccggctctg actgaccgcg
     2641 ttactcccac aggtgagcgg gcgggacggc ccttctcctc cgggctgtaa ttagcgcttg
     2701 gtttaatgac ggcttgtttc ttttctgtgg ctgcgtgaaa gccttgaggg gctccgggag
     2761 ggccctttgt gcggggggag cggctcgggg ctgtccgcgg ggggacggct gccttcgggg
     2821 gggacggggc agggcggggt tcggcttctg gcgtgtgacc ggcggctcta gagcctctgc
     2881 taaccatgtt catgccttct tctttttcct acagctcctg ggcaacgtgc tggttattgt
     2941 gctgtctcat cattttggca aagaattgga tccaccggtc gccaccatgg tgagcaaggg
     3001 cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg
     3061 ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct
     3121 gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccacctt
     3181 cggctacggc ctgcagtgct tcgcccgcta ccccgaccac atgaagcagc acgacttctt
     3241 caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg
     3301 caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga
     3361 gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa
     3421 ctacaacagc cacaacgtct atatcatggc cgacaagcag aagaacggca tcaaggtgaa
     3481 cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc actaccagca
     3541 gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagctacca
     3601 gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt
     3661 gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagtaaagcg gccgcctcga
     3721 ctagggataa cagggtaatt gtttgaatga ggcttcagta ctttacagaa tcgttgcctg
     3781 cacatcttgg aaacacttgc tgggattact tcgacttctt aacccaacag aaggctcgag
     3841 aaggtatatt gctgttgaca gtgagcgccc gcctgaagtc tctgattaat agtgaagcca
     3901 cagatgtatt aatcagagac ttcaggcggt tgcctactgc ctcggacttc aaggggctag
     3961 aattcgagca attatcttgt ttactaaaac tgaatacctt gctatctctt tgatacattt
     4021 ttacaaagct gaattaaaat ggtataaatt aaatcacttt tttcaattga cgcgtaagct
     4081 tatcgataat caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta
     4141 tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc
     4201 ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga
     4261 ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac
     4321 ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc
     4381 cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc
     4441 tcggctgttg ggcactgaca attccgtggt gttgtcgggg aaatcatcgt cctttccttg
     4501 gctgctcgcc tgtgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc
     4561 ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc
     4621 gcgtcttcgc cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcatcg
     4681 ataccgtcga ctcgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt
     4741 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     4801 taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     4861 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg
     4921 gtgggctcta tggcttctga ggcggaaaga accagctggg gctcgactag agcatggcta
     4981 cgtagataag tagcatggcg ggttaatcat taactacaag gaacccctag tgatggagtt
     5041 ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg
     5101 acgcccgggc ggcctcagtg agcgagcgag cgcgcagagc tttttgcaaa agcctaggcc
     5161 tccaaaaaag cctcctcact acttctggaa tagctcagag gccgaggcgg cctcggcctc
     5221 tgcataaata aaaaaaatta gtcagccatg gggcggagaa tgggcggaac tgggcggagt
     5281 taggggcggg atgggcggag ttaggggcgg gactatggtt gctgactaat tgagatgcat
     5341 gctttgcata cttctgcctg ctggggagcc tggggacttt ccacacctgg ttgctgacta
     5401 attgagatgc atgctttgca tacttctgcc tgctggggag cctggggact ttccacaccc
     5461 taactgacac acattccaca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     5521 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     5581 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     5641 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     5701 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     5761 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     5821 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     5881 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     5941 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     6001 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     6061 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     6121 ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct
     6181 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     6241 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     6301 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca
     6361 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat
     6421 ta
//
