LOCUS       sequence-134367-        3232 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cloning vector for expression of sgRNA-1xPBSb.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3232)
  AUTHORS   Cheng AW, Jillette N, Lee P, Plaskon D, Fujiwara Y, Wang W, 
            Taghbalout A, Wang H
  TITLE     Casilio: a versatile CRISPR-Cas9-Pumilio hybrid for gene regulation 
            and genomic labeling.
  JOURNAL   Cell Res. 2016 Feb;26(2):254-7. doi: 10.1038/cr.2016.3. Epub 2016 
            Jan 15.
  PUBMED    26768771
REFERENCE   2  (bases 1 to 3232)
  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/cr.2016.3";
            journalName: "Cell Res"; date: "2016-02"; volume: "26"; issue: "2"; 
            pages: "254-7"
COMMENT     Sequence Label: pAC1422-pX-sgRNA-1xPBSb
FEATURES             Location/Qualifiers
     source          1..3232
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        9..257
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     9..29
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     180..199
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     repeat_region   495..635
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   495..624
                     /label=AAV2 ITR
     rep_origin      710..1165
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(797..816)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1007..1028
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1182..1201)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1301..1323
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1361..1379)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1447..1551
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1552..1620,1621..2412)
                     /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     complement(1770..1789)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2583..3171
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3072..3091
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ggcgcgccga gggcctattt cccatgattc cttcatattt gcatatacga tacaaggctg
       61 ttagagagat aattggaatt aatttgactg taaacacaaa gatattagta caaaatacgt
      121 gacgtagaaa gtaataattt cttgggtagt ttgcagtttt aaaattatgt tttaaaatgg
      181 actatcatat gcttaccgta acttgaaagt atttcgattt cttggcttta tatatcttgt
      241 ggaaaggacg aaacaccggg tcttcgagaa gacctgttta agagctatgc tggaaacagc
      301 atagcaagtt taaataaggc tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg
      361 ccaattgggt ctccagatgc cttgatatag ccagatcttt ttttgtttta gagctagaaa
      421 tagcaagtta aaataaggct agtccgtagc gcgtgcgcca attctgcaga caaatggcgg
      481 cgcgccgcgg ccgcaggaac ccctagtgat ggagttggcc actccctctc tgcgcgctcg
      541 ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc ccgggctttg cccgggcggc
      601 ctcagtgagc gagcgagcgc gcagctgcct gcaggggcgc ctgatgcggt attttctcct
      661 tacgcatctg tgcggtattt cacaccgcat acgtcaaagc aaccatagta cgcgccctgt
      721 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
      781 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
      841 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
      901 cacctcgacc ccaaaaaact tgatttgggt gatggttcac gtagtgggcc atcgccctga
      961 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     1021 caaactggaa caacactcaa ccctatctcg ggctattctt ttgatttata agggattttg
     1081 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
     1141 aacaaaatat taacgtttac aattttatgg tgcactctca gtacaatctg ctctgatgcc
     1201 gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
     1261 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
     1321 aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt
     1381 ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac ttttcgggga
     1441 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     1501 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
     1561 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     1621 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
     1681 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
     1741 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
     1801 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
     1861 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     1921 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
     1981 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     2041 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
     2101 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     2161 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     2221 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtggaag ccgcggtatc
     2281 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     2341 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     2401 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
     2461 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     2521 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     2581 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     2641 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     2701 ttcagcagag cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac
     2761 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     2821 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     2881 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     2941 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     3001 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     3061 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     3121 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     3181 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gt
//
