LOCUS       sequence-199149-        2962 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For cloning of guide RNAs compatible with PguCas13b. Contains a 3' 
            direct repeat. Clone using BbsI (BpiI). F overhang cacc. R overhang 
            caac.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2962)
  AUTHORS   Cox DBT, Gootenberg JS, Abudayyeh OO, Franklin B, Kellner MJ, Joung 
            J, Zhang F
  TITLE     RNA editing with CRISPR-Cas13.
  JOURNAL   Science. 2017 Oct 25. pii: eaaq0180. doi: 10.1126/science.aaq0180.
  PUBMED    29070703
REFERENCE   2  (bases 1 to 2962)
  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: "Science. 
            2017 Oct 25. pii: eaaq0180. doi: 10.1126/science.aaq0180."
COMMENT     Sequence Label: pC0042-PguCas13b crRNA backbone
FEATURES             Location/Qualifiers
     source          1..2962
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(74..865,866..934))
                     /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     697..716
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(935..1039)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1107..1125
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1163..1185)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     1285..1304
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1498..1520
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1512..1529
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1513..1529
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             1572..1583
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     promoter        complement(1613..1861)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(1671..1690)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(1841..1861)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(1875..1891)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1875..1891)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1888..1910)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1899..1915
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(1923..1929,1930..1947,1948..1953))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1968..1989
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2106..2123)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2277..2865)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2357..2376)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
       61 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
      121 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
      181 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
      241 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
      301 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
      361 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
      421 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
      481 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
      541 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
      601 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
      661 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
      721 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
      781 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
      841 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
      901 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
      961 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     1021 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca
     1081 ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtctcgcgc
     1141 gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt
     1201 gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg
     1261 ggtgtcgggg ctggcttaac tatgcggcat cagagcagat tgtactgaga gtgcaccata
     1321 tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg cgccattcgc
     1381 cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc
     1441 agctggcgaa agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc
     1501 agtcacgacg ttgtaaaacg acggccagtg aattcgagct cggtaccaaa aaagttgtgt
     1561 gtacccttca aatagagggt agatccaact tgtcttcgtc ccaggaagac atggtgtttc
     1621 gtcctttcca caagatatat aaagccaaga aatcgaaata ctttcaagtt acggtaagca
     1681 tatgatagtc cattttaaaa cataatttta aaactgcaaa ctacccaaga aattattact
     1741 ttctacgtca cgtattttgt actaatatct ttgtgtttac agtcaaatta attccaatta
     1801 tctctctaac agccttgtat cgtatatgca aatatgaagg aatcatggga aataggccct
     1861 ctggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     1921 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     1981 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     2041 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     2101 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     2161 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     2221 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     2281 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     2341 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     2401 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     2461 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     2521 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     2581 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     2641 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     2701 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     2761 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     2821 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     2881 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     2941 agattatcaa aaaggatctt ca
//
