LOCUS       sequence-209270-        5211 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector expressing non-targeting AsCpf1 crRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5211)
  AUTHORS   Tekin H, Simmons S, Cummings B, Gao L, Adiconis X, Hession CC, 
            Ghoshal A, Dionne D, Choudhury SR, Yesilyurt V, Sanjana NE, Shi X, 
            Lu C, Heidenreich M, Pan JQ, Levin JZ, Zhang F
  TITLE     Effects of 3D culturing conditions on the transcriptomic profile of 
            stem-cell-derived neurons.
  JOURNAL   Nat Biomed Eng. 2018 Jul;2(7):540-554. doi: 
            10.1038/s41551-018-0219-9. Epub 2018 Apr 9.
  PUBMED    30271673
REFERENCE   2  (bases 1 to 5211)
  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: 
            "10.1038/s41551-018-0219-9"; journalName: "Nat Biomed Eng"; date: 
            "2018-07"; volume: "2"; issue: "7"; pages: "540-554"
COMMENT     Sequence Label: pU6-crRNA(Non-targeting)
FEATURES             Location/Qualifiers
     source          1..5211
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     promoter        158..406
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     158..178
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     329..348
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     promoter        494..941
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     CDS             979..1683
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(1121..1139)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1389..1408
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    2000..2476
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2199..2218)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   2485..2614
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(2676..3264)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2756..2775)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3435..4226,4227..4295))
                     /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     4058..4077
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4296..4400)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4468..4486
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4524..4546)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4646..4665
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(4682..5137)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4819..4840)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5031..5050
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgcg ctagcctgag ggcctatttc ccatgattcc
      181 ttcatatttg catatacgat acaaggctgt tagagagata attggaatta atttgactgt
      241 aaacacaaag atattagtac aaaatacgtg acgtagaaag taataatttc ttgggtagtt
      301 tgcagtttta aaattatgtt ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta
      361 tttcgatttc ttggctttat atatcttgtg gaaaggacga aacaccgtaa tttctactct
      421 tgtagattat ctgtgtagcc ggtgtggtag tttttcgacg cgtgtgtcta gactgcagag
      481 ggccctgcgt atgagtgcaa gtgggtttta ggaccaggat gaggcggggt gggggtgcct
      541 acctgacgac cgaccccgac ccactggaca agcacccaac ccccattccc caaattgcgc
      601 atcccctatc agagaggggg aggggaaaca ggatgcggcg aggcgcgtgc gcactgccag
      661 cttcagcacc gcggacagtg ccttcgcccc cgcctggcgg cgcgcgccac cgccgcctca
      721 gcactgaagg cgcgctgacg tcactcgccg gtcccccgca aactcccctt cccggccacc
      781 ttggtcgcgt ccgcgccgcc gccggcccag ccggaccgca ccacgcgagg cgcgagatag
      841 gggggcacgg gcgcgaccat ctgcgctgcg gcgccggcga ctcagcgctg cctcagtctg
      901 cggtgggcag cggaggagtc gtgtcgtgcc tgagagcgca gtcgagaagg taccggatcc
      961 tctagagtcg acgccaccgt gagcaagggc gaggaggata acatggccat catcaaggag
     1021 ttcatgcgct tcaaggtgca catggagggc tccgtgaacg gccacgagtt cgagatcgag
     1081 ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa ggtgaccaag
     1141 ggtggccccc tgcccttcgc ctgggacatc ctgtcccctc agttcatgta cggctccaag
     1201 gcctacgtga agcaccccgc cgacatcccc gactacttga agctgtcctt ccccgagggc
     1261 ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg tggtgaccgt gacccaggac
     1321 tcctccctgc aggacggcga gttcatctac aaggtgaagc tgcgcggcac caacttcccc
     1381 tccgacggcc ccgtaatgca gaagaagacc atgggctggg aggcctcctc cgagcggatg
     1441 taccccgagg acggcgccct gaagggcgag atcaagcaga ggctgaagct gaaggacggc
     1501 ggccactacg acgctgaggt caagaccacc tacaaggcca agaagcccgt gcagctgccc
     1561 ggcgcctaca acgtcaacat caagttggac atcacctccc acaacgagga ctacaccatc
     1621 gtggaacagt acgaacgcgc cgagggccgc cactccaccg gcggcatgga cgagctgtac
     1681 aagtccggac tcagatctcg agaggaggag gaggagacag acagcaggat gccccacctc
     1741 gacagccccg gcagctccca gccgagacgc tccttcctct caagggtgat cagggcagcg
     1801 ctaccgttgc agctgcttct gctgctgctg ctgctcctgg cctgcctgct acctgcctct
     1861 gaagatgact acagctgcac ccaggccaac aactttgccc gatccttcta ccccatgctg
     1921 cggtacacca acgggccacc tcccacctag gaattcgata tcaagcttat cgataccgag
     1981 cgctgctcga gagatctacg ggtggcatcc ctgtgacccc tccccagtgc ctctcctggc
     2041 cctggaagtt gccactccag tgcccaccag ccttgtccta ataaaattaa gttgcatcat
     2101 tttgtctgac taggtgtcct tctataatat tatggggtgg aggggggtgg tatggagcaa
     2161 ggggcaagtt gggaagacaa cctgtagggc ctgcggggtc tattgggaac caagctggag
     2221 tgcagtggca caatcttggc tcactgcaat ctccgcctcc tgggttcaag cgattctcct
     2281 gcctcagcct cccgagttgt tgggattcca ggcatgcatg accaggctca gctaattttt
     2341 gtttttttgg tagagacggg gtttcaccat attggccagg ctggtctcca actcctaatc
     2401 tcaggtgatc tacccacctt ggcctcccaa attgctggga ttacaggcgt gaaccactgc
     2461 tcccttccct gtccttgcgg ccgcaggaac ccctagtgat ggagttggcc actccctctc
     2521 tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc ccgggcggcc
     2581 tcagtgagcg agcgagcgcg cagctgcctg caggacatgt gagcaaaagg ccagcaaaag
     2641 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     2701 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     2761 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     2821 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     2881 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     2941 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     3001 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     3061 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     3121 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     3181 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     3241 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     3301 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     3361 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     3421 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     3481 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     3541 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     3601 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     3661 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     3721 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     3781 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     3841 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     3901 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     3961 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     4021 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     4081 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     4141 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     4201 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     4261 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     4321 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     4381 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc
     4441 attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg
     4501 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct
     4561 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc
     4621 gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat
     4681 aaaattgtaa acgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca
     4741 ttttttaacc aatagaccga aatcggcaaa atcccttata aatcaaaaga atagcccgag
     4801 atagagttga gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc
     4861 aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc
     4921 aaatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc
     4981 ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa
     5041 gcgaaaggag cgggcgctaa ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc
     5101 acacccgccg cgcttaatgc gccgctacag ggcgcgtact atggttgctt tgacgtatgc
     5161 ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg catcaggcgc c
//
