LOCUS       sequence-134366-        3237 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cloning vector for expression of sgRNA-2xPBSb.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3237)
  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 3237)
  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: pAC1423-pX-sgRNA-2xPBSb
FEATURES             Location/Qualifiers
     source          1..3237
                     /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   500..640
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   500..629
                     /label=AAV2 ITR
     rep_origin      715..1170
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(802..821)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1012..1033
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1187..1206)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1306..1328
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1366..1384)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1452..1556
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1557..1625,1626..2417)
                     /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(1775..1794)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2588..3176
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3077..3096
                     /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 ctccagattt gatatagcct tgatataaga tctttttttg ttttagagct
      421 agaaatagca agttaaaata aggctagtcc gtagcgcgtg cgccaattct gcagacaaat
      481 ggcggcgcgc cgcggccgca ggaaccccta gtgatggagt tggccactcc ctctctgcgc
      541 gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg
      601 gcggcctcag tgagcgagcg agcgcgcagc tgcctgcagg ggcgcctgat gcggtatttt
      661 ctccttacgc atctgtgcgg tatttcacac cgcatacgtc aaagcaacca tagtacgcgc
      721 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
      781 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
      841 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
      901 tacggcacct cgaccccaaa aaacttgatt tgggtgatgg ttcacgtagt gggccatcgc
      961 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     1021 tgttccaaac tggaacaaca ctcaacccta tctcgggcta ttcttttgat ttataaggga
     1081 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     1141 attttaacaa aatattaacg tttacaattt tatggtgcac tctcagtaca atctgctctg
     1201 atgccgcata gttaagccag ccccgacacc cgccaacacc cgctgacgcg ccctgacggg
     1261 cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt
     1321 gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc gtgatacgcc
     1381 tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc
     1441 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     1501 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     1561 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     1621 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     1681 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     1741 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
     1801 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     1861 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     1921 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     1981 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     2041 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     2101 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     2161 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     2221 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt ggaagccgcg
     2281 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     2341 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     2401 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     2461 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     2521 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     2581 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     2641 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     2701 ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc
     2761 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     2821 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     2881 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     2941 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     3001 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     3061 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     3121 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     3181 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgt
//
