LOCUS       sequence-134365-        3326 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cloning vector for expression of sgRNA-10xPBSb.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3326)
  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 3326)
  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: pAC1424-pX-sgRNA-10xPBSb
FEATURES             Location/Qualifiers
     source          1..3326
                     /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   589..729
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   589..718
                     /label=AAV2 ITR
     rep_origin      804..1259
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(891..910)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1101..1122
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1276..1295)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1395..1417
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1455..1473)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1541..1645
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1646..1714,1715..2506)
                     /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(1864..1883)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2677..3265
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3166..3185
                     /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 tgatatagcc ttgatatagc cttgatatag
      421 ccttgatata agatttgata tagccttgat atagccttga tatagccttg atatagcctt
      481 gatataagat ctttttttgt tttagagcta gaaatagcaa gttaaaataa ggctagtccg
      541 tagcgcgtgc gccaattctg cagacaaatg gcggcgcgcc gcggccgcag gaacccctag
      601 tgatggagtt ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa
      661 aggtcgcccg acgcccgggc tttgcccggg cggcctcagt gagcgagcga gcgcgcagct
      721 gcctgcaggg gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
      781 gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
      841 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
      901 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
      961 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgattt
     1021 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     1081 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     1141 ctcgggctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     1201 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt
     1261 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc
     1321 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     1381 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     1441 cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat
     1501 ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     1561 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     1621 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     1681 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     1741 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     1801 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     1861 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
     1921 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     1981 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     2041 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     2101 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     2161 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
     2221 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     2281 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     2341 atctggagcc ggtgagcgtg gaagccgcgg tatcattgca gcactggggc cagatggtaa
     2401 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     2461 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     2521 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     2581 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     2641 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     2701 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     2761 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     2821 tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     2881 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     2941 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     3001 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     3061 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     3121 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     3181 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     3241 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     3301 cttttgctgg ccttttgctc acatgt
//
