LOCUS       sequence-211313-        2615 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  U6-driven sgRNA expression plasmid for Streptococcus thermophilus 
            CRISPR1 (St1Cas9 LMD-9)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2615)
  AUTHORS   Agudelo D, Carter S, Velimirovic M, Duringer A, Rivest JF, Levesque 
            S, Loehr J, Mouchiroud M, Cyr D, Waters PJ, Laplante M, Moineau S, 
            Goulet A, Doyon Y
  TITLE     Versatile and robust genome editing with Streptococcus thermophilus 
            CRISPR1-Cas9.
  JOURNAL   Genome Res. 2020 Jan;30(1):107-117. doi: 10.1101/gr.255414.119. Epub
            2020 Jan 3.
  PUBMED    31900288
REFERENCE   2  (bases 1 to 2615)
  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: 
            "10.1101/gr.255414.119"; journalName: "Genome Res"; date: "2020-01";
            volume: "30"; issue: "1"; pages: "107-117"
COMMENT     Sequence Label: St1Cas9_LMD-9_sgRNA_pUC19
FEATURES             Location/Qualifiers
     source          1..2615
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        56..304
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     56..76
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     227..246
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(438..454)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(438..454)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(451..473)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    462..478
                     /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(486..492,493..510,511..516))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    531..552
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(669..686)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(840..1428)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(920..939)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1599..2390,2391..2459))
                     /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     2222..2241
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2460..2564)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggctttaaag gaaccaattc agtcgactgg atccggtacc aaggtcgggc aggaagaggg
       61 cctatttccc atgattcctt catatttgca tatacgatac aaggctgtta gagagataat
      121 tagaattaat ttgactgtaa acacaaagat attagtacaa aatacgtgac gtagaaagta
      181 ataatttctt gggtagtttg cagttttaaa attatgtttt aaaatggact atcatatgct
      241 taccgtaact tgaaagtatt tcgatttctt ggctttatat atcttgtgga aaggacgaaa
      301 cacccgagac gattaatgcg tctcggtctt tgtactctgg taccagaagc tacaaagata
      361 aggcttcatg ccgaaatcaa caccctgtca ttttatggca gggtgttttt tttactagta
      421 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
      481 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
      541 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
      601 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
      661 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
      721 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
      781 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
      841 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
      901 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
      961 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     1021 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     1081 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     1141 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     1201 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     1261 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     1321 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     1381 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     1441 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     1501 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     1561 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     1621 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     1681 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     1741 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     1801 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     1861 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     1921 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     1981 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     2041 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     2101 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     2161 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     2221 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     2281 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     2341 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     2401 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     2461 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     2521 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     2581 gtgccacctg acgtcgctag ctgtacaaaa aagca
//
