LOCUS       pHnS_KI;Pdha1_Donor;3xV5_KO;Template        4802 bp    DNA     circular UNA 11-JUL-2025
DEFINITION  pAAV-U6sgRNA(SapI)_hSyn-GFP-KASH-bGH (SpGuide acceptor). AAV
            plasmid for sgRNA cloning. GFP-KASH fusion facilitates FACS sorting
            of cells and nuclei. .
ACCESSION   urn.local...24-jeblee4
VERSION     urn.local...24-jeblee4
KEYWORDS    .
SOURCE      
  ORGANISM  .
FEATURES             Location/Qualifiers
     source          1..4802
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /note="inverted terminal repeat of adeno-associated virus
                     serotype 2"
                     /standard_name="AAV2 ITR"
     repeat_region   12..141
                     /standard_name="AAV2 ITR"
     misc_feature    153..171
                     /standard_name="Primer site #1"
     misc_feature    172..461
                     /standard_name="bU6"
     misc_feature    462..481
                     /standard_name=1
     misc_feature    464..469
                     /standard_name="BbsI"
     misc_feature    482..567
                     /standard_name="Scaffold cr1"
     misc_feature    568..574
                     /standard_name="polyT"
     misc_feature    575..594
                     /standard_name="Primer site #2"
     misc_feature    595..908
                     /standard_name="mU6"
     protein_bind    789..822
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core
                     sequence (GCATACAT)."
                     /standard_name="loxP"
     primer_bind     826..846
                     /standard_name="mU6-F"
     primer_bind     complement(887..908)
                     /standard_name="SEQ_mU6_last_R"
     misc_feature    909..928
                     /standard_name=2
     misc_feature    929..1016
                     /standard_name="Scaffold cr2"
     misc_feature    1017..1023
                     /standard_name="polyT"
     misc_feature    1024..1042
                     /standard_name="Primer site #3"
     source          1043..1052
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    1053..1319
                     /standard_name="hU6"
     primer_bind     1071..1091
                     /standard_name="hU6-F"
     misc_feature    1320..1339
                     /standard_name=3
     misc_feature    1332..1337
                     /standard_name="BbsI"
     misc_feature    1340..1427
                     /standard_name="Scaffold cr3"
     misc_feature    1428..1434
                     /standard_name="polyT"
     misc_feature    1435..1456
                     /standard_name="Primer site #4"
     regulatory      1466..1680
                     /note="Geneious type: promoter"
                     /note="human H1 RNA promoter"
                     /standard_name="H1 promoter"
     primer_bind     1601..1620
                     /standard_name="H1"
     misc_feature    1686..1705
                     /standard_name="rPdha1"
     misc_feature    1706..1793
                     /standard_name="Scaffold cr4"
     misc_feature    1794..1800
                     /standard_name="polyT"
     misc_feature    complement(1817..1839)
                     /standard_name="rPdha1"
     CDS             1848..1889
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /transl_table=1
                     /translation="GKPIPNPLLGLDST"
                     /standard_name="V5 tag"
     CDS             1899..1940
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /transl_table=1
                     /translation="GKPIPNPLLGLDST"
                     /standard_name="V5 tag"
     CDS             1950..1991
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /transl_table=1
                     /translation="GKPIPNPLLGLDST"
                     /standard_name="V5 tag"
     misc_feature    1992..2006
                     /locus_tag="X"
                     /standard_name="X"
     misc_feature    complement(2014..2036)
                     /standard_name="rPdha1"
     repeat_region   2065..2205
                     /note="inverted terminal repeat of adeno-associated virus
                     serotype 2"
                     /standard_name="AAV2 ITR"
     repeat_region   2065..2194
                     /standard_name="AAV2 ITR"
     rep_origin      2280..2735
     primer_bind     complement(2367..2386)
                     /standard_name="F1ori-R"
     primer_bind     2577..2598
                     /standard_name="F1ori-F"
     primer_bind     complement(2752..2771)
                     /standard_name="pRS-marker"
     primer_bind     2871..2893
                     /standard_name="pGEX 3'"
     primer_bind     complement(2931..2949)
                     /standard_name="pBRforEco"
     regulatory      3017..3121
                     /note="Geneious type: promoter"
                     /gene="bla"
                     /standard_name="AmpR promoter"
     CDS             join(3122..3190,3191..3982)
                     /codon_start=1
                     /gene="bla"
                     /note="confers resistance to ampicillin, carbenicillin,
                     and related antibiotics"
                     /product="beta-lactamase"
                     /transl_table=1
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY
                     IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVE
                     YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL
                     DRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL
                     LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA
                     EIGASLIKHW*"
                     /standard_name="AmpR"
     primer_bind     complement(3340..3359)
                     /standard_name="Amp-R"
     rep_origin      4153..4741
     primer_bind     4642..4661
                     /standard_name="pBR322ori-F"
ORIGIN      
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgcc ggtaggtccc tcgacgttaa ccgtgaccga
      181 gcttgtctgc ccgcgcagtc cactagacag acgcgcagag gtcgggcggg cctagggcca
      241 atttcccatg gttcccttca tttgcatata tgatgtaata aggttatgga gactattaaa
      301 cttagcccta atcaaactat atgatgataa tgtgtgtggt acaaaaggtc ataacttatt
      361 atatactttg aaacttaaaa aagggttaca gtttagtcac cataactgta aaattttttc
      421 tattcttagc tttatatagt tcttgagagg ccatgtttat gctgtcttca tcatggccga
      481 cgtttaagag ctaagctgga aacagcatag caagtttaaa taaggctagt ccgttatcaa
      541 cttgaaaaag tggcaccgag tcggtgcttt ttttccagtc tgactatcac gatggatccg
      601 acgcgccatc tctaggcccg cgccggcccc ctcgcacgga cttgtgggag aagctcggct
      661 actcccctgc cccggttaat ttgcatataa tatttcctag taactataga ggcttaatgt
      721 gcgataaaag acagataatc tgttcttttt aatactagct acattttaca tgataggctt
      781 ggatttctat aacttcgtat agcatacatt atacgaagtt ataaacagca caaaaggaaa
      841 ctcaccctaa ctgtaaagta attgtgtgtt ttgagactat aagtatccct tggagaacca
      901 ccttgttggt tcgcattatc cgaaccatgt ttgagagcta agcagaaagc tgcatagcaa
      961 gttcaaataa ggctagtccg tacacaactt gaaaaagtgg cagccgagtc ggctgctttt
     1021 tttgctcgaa tctacactca gctatggcgc gccccaaggt cgggcaggaa gagggcctat
     1081 ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag ataattggaa
     1141 ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga aagtaataat
     1201 ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat atgcttaccg
     1261 taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagcc agaaacatgt
     1321 aagcgtcgca agaagacggg tttcagagct aagcacaaga gtgcatagca agttgaaata
     1381 aggctagtcc gtttacaact tgaaaaagtg gcacccgagt cgggtgcttt ttttctcgag
     1441 tactaggatc cattaggcgg tctaggaacg ctgacgtcat caacccgctc caaggaatcg
     1501 cgggcccagt gtcactaggc gggaacaccc agcgcgcgtg cgccctggca ggaagatggc
     1561 tgtgagggac aggggagtgg cgccctgcaa tatttgcatg tcgctatgtg ttctgggaaa
     1621 tcaccataaa cgtgaaatgt ctttggattt gggaatctta taagttctgt atgagaccac
     1681 caccggaccc acccttcgaa gtgcggttta agagctaagc tcagaagagc atagcaagtt
     1741 taaataaggc tagtccgtac tcaacttgaa aaagtggcag ccgagtcggc tgcttttttt
     1801 ggtctgagac gccgcgccac gcacttcgaa gggtgggtcc tcgagaagga aaacctatac
     1861 cgaaccccct ccttggactt gatagtactc agcagcaggg taagcccatt ccaaacccac
     1921 ttctcggtct ggatagcaca cagcaacaag gtaagcctat ccctaaccct ctgttgggcc
     1981 tggactccac gctagctagc tagctaggga tccccacgca cttcgaaggg tgggtccacg
     2041 cgtgtgtcta gactgcgcgg ccgcaggaac ccctagtgat ggagttggcc actccctctc
     2101 tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc ccgggctttg
     2161 cccgggcggc ctcagtgagc gagcgagcgc gcagctgcct gcaggggcgc ctgatgcggt
     2221 attttctcct tacgcatctg tgcggtattt cacaccgcat acgtcaaagc aaccatagta
     2281 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     2341 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     2401 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     2461 tgctttacgg cacctcgacc ccaaaaaact tgatttgggt gatggttcac gtagtgggcc
     2521 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     2581 actcttgttc caaactggaa caacactcaa ccctatctcg ggctattctt ttgatttata
     2641 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     2701 cgcgaatttt aacaaaatat taacgtttac aattttatgg tgcactctca gtacaatctg
     2761 ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg
     2821 acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg
     2881 catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat
     2941 acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac
     3001 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     3061 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
     3121 tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc
     3181 tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc
     3241 acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc
     3301 cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc
     3361 ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt
     3421 ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt
     3481 atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat
     3541 cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct
     3601 tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat
     3661 gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc
     3721 ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg
     3781 ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc
     3841 tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta
     3901 cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc
     3961 ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga
     4021 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
     4081 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     4141 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     4201 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     4261 ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt agccgtagtt
     4321 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     4381 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     4441 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     4501 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     4561 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     4621 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     4681 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     4741 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     4801 gt
//
