LOCUS       sequence-412377-        5807 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Control plasmid expresses mCherry under the hSyn1 promoter with 
            sgRNAs against LacZ and GFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5807)
  AUTHORS   Duncan GJ, Ingram SD, Emberley K, Hill J, Cordano C, Abdelhak A, 
            McCane M, Jenks JE, Jabassini N, Ananth K, Ferrara SJ, Stedelin B, 
            Sivyer B, Aicher SA, Scanlan TS, Watkins TA, Mishra A, Nelson JW, 
            Green AJ, Emery B
  TITLE     Remyelination protects neurons from DLK-mediated neurodegeneration.
  JOURNAL   Nat Commun. 2024 Oct 23;15(1):9148. doi: 10.1038/s41467-024-53429-5.
  PUBMED    39443516
REFERENCE   2  (bases 1 to 5807)
  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"; journalName: "Nat 
            Commun."; date: "2024-10-23"; volume: "15(1):9148. doi"; pages: " 
            10.1038/s41467-024-53429-5"
COMMENT     Sequence Label: AAV-U6-sgLacZ-U6-sgGFP-hSyn1-mCherry
FEATURES             Location/Qualifiers
     source          1..5807
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(96..115)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      909..1497
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1398..1417
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   1559..1688
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        1720..1968
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        1988..2063
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        2154..2402
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        2423..2498
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        2606..3053
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     CDS             3085..3795
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(3230..3248)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3498..3517
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    4112..4588
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4311..4330)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4628..4768
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4628..4757
                     /label=AAV2 ITR
     rep_origin      4843..5298
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4930..4949)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5140..5161
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5315..5334)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5434..5456
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5494..5512)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5580..5684
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5685..5753,5754..5807,1..738)
                     /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"
ORIGIN
        1 gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa
       61 gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt
      121 attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt
      181 gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc
      241 agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga
      301 ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat
      361 cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct
      421 gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc
      481 cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg
      541 gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc
      601 ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg
      661 acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca
      721 ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta gattgattta
      781 aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc
      841 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
      901 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
      961 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     1021 actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc
     1081 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     1141 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     1201 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     1261 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     1321 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     1381 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     1441 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     1501 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgtcc
     1561 tgcaggcagc tgcgcgctcg ctcgctcact gaggccgccc gggcgtcggg cgacctttgg
     1621 tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga gtggccaact ccatcactag
     1681 gggttcctgc ggccgcacgc gttcgagaat ggcgcatgtg agggcctatt tcccatgatt
     1741 ccttcatatt tgcatatacg atacaaggct gttagagaga taattggaat taatttgact
     1801 gtaaacacaa agatattagt acaaaatacg tgacgtagaa agtaataatt tcttgggtag
     1861 tttgcagttt taaaattatg ttttaaaatg gactatcata tgcttaccgt aacttgaaag
     1921 tatttcgatt tcttggcttt atatatcttg tggaaaggac gaaacacctg cgatacgccc
     1981 acgcgatgtt ttagagctag aaatagcaag ttaaaataag gctagtccgt tatcaacttg
     2041 aaaaagtggc accgagtcgg tgcttttttg ttttagagct agaaatagca agttaaaata
     2101 aggctagtcc gtttttagcg cgtgcgccaa ttctgcagac aaatggctct agagagggcc
     2161 tatttcccat gattccttca tatttgcata tacgatacaa ggctgttaga gagataattg
     2221 gaattaattt gactgtaaac acaaagatat tagtacaaaa tacgtgacgt agaaagtaat
     2281 aatttcttgg gtagtttgca gttttaaaat tatgttttaa aatggactat catatgctta
     2341 ccgtaacttg aaagtatttc gatttcttgg ctttatatat cttgtggaaa ggacgaaaca
     2401 ccgccgtcca gctcgaccag gagttttaga gctagaaata gcaagttaaa ataaggctag
     2461 tccgttatca acttgaaaaa gtggcaccga gtcggtgctt ttttgtttta gagctagaaa
     2521 tagcaagtta aaataaggct agtccgtttt tagcgcgtgc gccaattctg cagacaaatg
     2581 gggtacaaat tgggccctgc gtatgagtgc aagtgggttt taggaccagg atgaggcggg
     2641 gtgggggtgc ctacctgacg accgaccccg acccactgga caagcaccca acccccattc
     2701 cccaaattgc gcatccccta tcagagaggg ggaggggaaa caggatgcgg cgaggcgcgt
     2761 gcgcactgcc agcttcagca ccgcggacag tgccttcgcc cccgcctggc ggcgcgcgcc
     2821 accgccgcct cagcactgaa ggcgcgctga cgtcactcgc cggtcccccg caaactcccc
     2881 ttcccggcca ccttggtcgc gtccgcgccg ccgccggccc agccggaccg caccacgcga
     2941 ggcgcgagat aggggggcac gggcgcgacc atctgcgctg cggcgccggc gactcagcgc
     3001 tgcctcagtc tgcggtgggc agcggaggag tcgtgtcgtg cctgagagcg cagtcgagaa
     3061 ggtaccggat cctctagagt cgacatggtg agcaagggcg aggaggataa catggccatc
     3121 atcaaggagt tcatgcgctt caaggtgcac atggagggct ccgtgaacgg ccacgagttc
     3181 gagatcgagg gcgagggcga gggccgcccc tacgagggca cccagaccgc caagctgaag
     3241 gtgaccaagg gtggccccct gcccttcgcc tgggacatcc tgtcccctca gttcatgtac
     3301 ggctccaagg cctacgtgaa gcaccccgcc gacatccccg actacttgaa gctgtccttc
     3361 cccgagggct tcaagtggga gcgcgtgatg aacttcgagg acggcggcgt ggtgaccgtg
     3421 acccaggact cctccctgca ggacggcgag ttcatctaca aggtgaagct gcgcggcacc
     3481 aacttcccct ccgacggccc cgtaatgcag aagaagacca tgggctggga ggcctcctcc
     3541 gagcggatgt accccgagga cggcgccctg aagggcgaga tcaagcagag gctgaagctg
     3601 aaggacggcg gccactacga cgctgaggtc aagaccacct acaaggccaa gaagcccgtg
     3661 cagctgcccg gcgcctacaa cgtcaacatc aagttggaca tcacctccca caacgaggac
     3721 tacaccatcg tggaacagta cgaacgcgcc gagggccgcc actccaccgg cggcatggac
     3781 gagctgtaca agtagtccgg actcagatct cgagaggagg aggaggagac agacagcagg
     3841 atgccccacc tcgacagccc cggcagctcc cagccgagac gctccttcct ctcaagggtg
     3901 atcagggcag cgctaccgtt gcagctgctt ctgctgctgc tgctgctcct ggcctgcctg
     3961 ctacctgcct ctgaagatga ctacagctgc acccaggcca acaactttgc ccgatccttc
     4021 taccccatgc tgcggtacac caacgggcca cctcccacct aggaattcga tatcaagctt
     4081 atcgataccg agcgctgctc gagagatcta cgggtggcat ccctgtgacc cctccccagt
     4141 gcctctcctg gccctggaag ttgccactcc agtgcccacc agccttgtcc taataaaatt
     4201 aagttgcatc attttgtctg actaggtgtc cttctataat attatggggt ggaggggggt
     4261 ggtatggagc aaggggcaag ttgggaagac aacctgtagg gcctgcgggg tctattggga
     4321 accaagctgg agtgcagtgg cacaatcttg gctcactgca atctccgcct cctgggttca
     4381 agcgattctc ctgcctcagc ctcccgagtt gttgggattc caggcatgca tgaccaggct
     4441 cagctaattt ttgttttttt ggtagagacg gggtttcacc atattggcca ggctggtctc
     4501 caactcctaa tctcaggtga tctacccacc ttggcctccc aaattgctgg gattacaggc
     4561 gtgaaccact gctcccttcc ctgtccttct gattttgtag gtaaccacgt gcggaccgag
     4621 cggccgcagg aacccctagt gatggagttg gccactccct ctctgcgcgc tcgctcgctc
     4681 actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg
     4741 agcgagcgag cgcgcagctg cctgcagggg cgcctgatgc ggtattttct ccttacgcat
     4801 ctgtgcggta tttcacaccg catacgtcaa agcaaccata gtacgcgccc tgtagcggcg
     4861 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgcct
     4921 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     4981 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     5041 accccaaaaa acttgatttg ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     5101 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     5161 gaacaacact caactctatc tcgggctatt cttttgattt ataagggatt ttgccgattt
     5221 cggtctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa
     5281 tattaacgtt tacaatttta tggtgcactc tcagtacaat ctgctctgat gccgcatagt
     5341 taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     5401 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     5461 caccgtcatc accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg
     5521 ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc
     5581 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     5641 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt
     5701 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     5761 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacga
//
