LOCUS       pHnS_KI;Sptbn4_Donor;3xV5_KO;Template        4789 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...2h-jebleeg
VERSION     urn.local...2h-jebleeg
KEYWORDS    .
SOURCE      
  ORGANISM  .
FEATURES             Location/Qualifiers
     source          1..4789
                     /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="mrb4sp#1"
     misc_feature    1706..1793
                     /standard_name="Scaffold cr4"
     misc_feature    1794..1800
                     /standard_name="polyT"
     misc_feature    complement(1817..1839)
                     /locus_tag="DRSR2(2)"
                     /standard_name="mrb4sp#1"
     CDS             1847..1888
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /transl_table=1
                     /translation="GKPIPNPLLGLDST"
                     /standard_name="V5 tag"
     CDS             1898..1939
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /transl_table=1
                     /translation="GKPIPNPLLGLDST"
                     /standard_name="V5 tag"
     CDS             1949..1990
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /transl_table=1
                     /translation="GKPIPNPLLGLDST"
                     /standard_name="V5 tag"
     misc_feature    complement(2001..2023)
                     /locus_tag="DRSR2(2)"
                     /standard_name="mrb4sp#1"
     repeat_region   2052..2192
                     /note="inverted terminal repeat of adeno-associated virus
                     serotype 2"
                     /standard_name="AAV2 ITR"
     repeat_region   2052..2181
                     /standard_name="AAV2 ITR"
     rep_origin      2267..2722
     primer_bind     complement(2354..2373)
                     /standard_name="F1ori-R"
     primer_bind     2564..2585
                     /standard_name="F1ori-F"
     primer_bind     complement(2739..2758)
                     /standard_name="pRS-marker"
     primer_bind     2858..2880
                     /standard_name="pGEX 3'"
     primer_bind     complement(2918..2936)
                     /standard_name="pBRforEco"
     regulatory      3004..3108
                     /note="Geneious type: promoter"
                     /gene="bla"
                     /standard_name="AmpR promoter"
     CDS             join(3109..3177,3178..3969)
                     /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(3327..3346)
                     /standard_name="Amp-R"
     rep_origin      4140..4728
     primer_bind     4629..4648
                     /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 caccgagcgt agggccagcg ggcgcgttta agagctaagc tcagaagagc atagcaagtt
     1741 taaataaggc tagtccgtac tcaacttgaa aaagtggcag ccgagtcggc tgcttttttt
     1801 ggtctgagac gccgcgccgg cgcccgctgg ccctacgctc tcgagaggaa aacctatacc
     1861 gaaccccctc cttggacttg atagtactca gcagcagggt aagcccattc caaacccact
     1921 tctcggtctg gatagcacac agcaacaagg taagcctatc cctaaccctc tgttgggcct
     1981 ggactccacg taggggatcc ccggcgcccg ctggccctac gctcacgcgt gtgtctagac
     2041 tgcgcggccg caggaacccc tagtgatgga gttggccact ccctctctgc gcgctcgctc
     2101 gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc
     2161 agtgagcgag cgagcgcgca gctgcctgca ggggcgcctg atgcggtatt ttctccttac
     2221 gcatctgtgc ggtatttcac accgcatacg tcaaagcaac catagtacgc gccctgtagc
     2281 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     2341 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     2401 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     2461 ctcgacccca aaaaacttga tttgggtgat ggttcacgta gtgggccatc gccctgatag
     2521 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     2581 actggaacaa cactcaaccc tatctcgggc tattcttttg atttataagg gattttgccg
     2641 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac
     2701 aaaatattaa cgtttacaat tttatggtgc actctcagta caatctgctc tgatgccgca
     2761 tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg
     2821 ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg
     2881 ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta
     2941 taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat
     3001 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     3061 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
     3121 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
     3181 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
     3241 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
     3301 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
     3361 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
     3421 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
     3481 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
     3541 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
     3601 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg
     3661 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
     3721 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
     3781 gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt
     3841 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
     3901 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
     3961 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
     4021 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     4081 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     4141 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     4201 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     4261 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
     4321 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     4381 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     4441 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     4501 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     4561 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     4621 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     4681 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     4741 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgt
//
