LOCUS       sequence-72014-e        3178 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  U6-driven sgRNA expressing cassette on a gateway donor vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3178)
  AUTHORS   Cheng AW, Wang H, Yang H, Shi L, Katz Y, Theunissen TW, Rangarajan 
            S, Shivalila CS, Dadon DB, Jaenisch R
  TITLE     Multiplexed activation of endogenous genes by CRISPR-on, an 
            RNA-guided transcriptional activator system.
  JOURNAL   Cell Res. 2013 Aug 27. doi: 10.1038/cr.2013.122.
  PUBMED    23979020
REFERENCE   2  (bases 1 to 3178)
  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: "Cell Res. 
            2013 Aug 27. doi: 10.1038/cr.2013.122."
COMMENT     Sequence Label: pAC155-pCR8-sgExpression
FEATURES             Location/Qualifiers
     source          1..3178
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     primer_bind     462..484
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     476..493
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     477..493
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    509..608
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     misc_RNA        complement(704..779)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(798..1046)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(856..875)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(1026..1046)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     protein_bind    complement(1057..1156)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1173..1192)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1174..1192)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1197..1213)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1197..1213)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             1644..2435
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      2528..3116
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3017..3036
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc caacgctagc atggatgttt tcccagtcac gacgttgtaa
      481 aacgacggcc agtcttaagc tcgggcccca aataatgatt ttattttgac tgatagtgac
      541 ctgttcgttg caacaaattg atgagcaatg cttttttata atgccaactt tgtacaaaaa
      601 agcaggctcc gcctctagag ccatttgtct gcagaattgg cgcacgcgct aaaaacggac
      661 tagccttatt ttaacttgct atttctagct ctaaaacaaa aaagcaccga ctcggtgcca
      721 ctttttcaag ttgataacgg actagcctta ttttaacttg ctatttctag ctctaaaaca
      781 ggtcttctcg aagacccggt gtttcgtcct ttccacaaga tatataaagc caagaaatcg
      841 aaatactttc aagttacggt aagcatatga tagtccattt taaaacataa ttttaaaact
      901 gcaaactacc caagaaatta ttactttcta cgtcacgtat tttgtactaa tatctttgtg
      961 tttacagtca aattaattcc aattatctct ctaacagcct tgtatcgtat atgcaaatat
     1021 gaaggaatca tgggaaatag gccctcacat gtggcgaccc agctttcttg tacaaagttg
     1081 gcattataaa aaataattgc tcatcaattt gttgcaacga acaggtcact atcagtcaaa
     1141 ataaaatcat tatttgccat ccagctgata tcccctatag tgagtcgtat tacatggtca
     1201 tagctgtttc ctggcagctc tggcccgtgt ctcaaaatct ctgatgttac attgcacaag
     1261 ataaaaatat atcatcatgc ctcctctaga ccagccagga cagaaatgcc tcgacttcgc
     1321 tgctgcccaa ggttgccggg tgacgcacac cgtggaaacg gatgaaggca cgaacccagt
     1381 ggacataagc ctgttcggtt cgtaagctgt aatgcaagta gcgtatgcgc tcacgcaact
     1441 ggtccagaac cttgaccgaa cgcagcggtg gtaacggcgc agtggcggtt ttcatggctt
     1501 gttatgactg tttttttggg gtacagtcta tgcctcgggc atccaagcag caagcgcgtt
     1561 acgccgtggg tcgatgtttg atgttatgga gcagcaacga tgttacgcag cagggcagtc
     1621 gccctaaaac aaagttaaac atcatgaggg aagcggtgat cgccgaagta tcgactcaac
     1681 tatcagaggt agttggcgtc atcgagcgcc atctcgaacc gacgttgctg gccgtacatt
     1741 tgtacggctc cgcagtggat ggcggcctga agccacacag tgatattgat ttgctggtta
     1801 cggtgaccgt aaggcttgat gaaacaacgc ggcgagcttt gatcaacgac cttttggaaa
     1861 cttcggcttc ccctggagag agcgagattc tccgcgctgt agaagtcacc attgttgtgc
     1921 acgacgacat cattccgtgg cgttatccag ctaagcgcga actgcaattt ggagaatggc
     1981 agcgcaatga cattcttgca ggtatcttcg agccagccac gatcgacatt gatctggcta
     2041 tcttgctgac aaaagcaaga gaacatagcg ttgccttggt aggtccagcg gcggaggaac
     2101 tctttgatcc ggttcctgaa caggatctat ttgaggcgct aaatgaaacc ttaacgctat
     2161 ggaactcgcc gcccgactgg gctggcgatg agcgaaatgt agtgcttacg ttgtcccgca
     2221 tttggtacag cgcagtaacc ggcaaaatcg cgccgaagga tgtcgctgcc gactgggcaa
     2281 tggagcgcct gccggcccag tatcagcccg tcatacttga agctagacag gcttatcttg
     2341 gacaagaaga agatcgcttg gcctcgcgcg cagatcagtt ggaagaattt gtccactacg
     2401 tgaaaggcga gatcaccaag gtagtcggca aataaccctc gagccaccca tgaccaaaat
     2461 cccttaacgt gagttacgcg tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     2521 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     2581 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     2641 ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc
     2701 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     2761 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     2821 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     2881 gaacgaccta caccgaactg agatacctac agcgtgagca ttgagaaagc gccacgcttc
     2941 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     3001 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     3061 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     3121 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgtt
//
