LOCUS       sequence-422138-        3244 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of tandem pre-sgRNA array pcr4 for LbCas12a.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3244)
  AUTHORS   Chen F, Lian M, Ma B, Gou S, Luo X, Yang K, Shi H, Xie J, Ge W, 
            Ouyang Z, Lai C, Li N, Zhang Q, Jin Q, Liang Y, Chen T, Wang J, Zhao
            X, Li L, Yu M, Ye Y, Wang K, Wu H, Lai L
  TITLE     Multiplexed base editing through Cas12a variant-mediated cytosine 
            and adenine base editors.
  JOURNAL   Commun Biol. 2022 Nov 2;5(1):1163. doi: 10.1038/s42003-022-04152-8.
  PUBMED    36323848
REFERENCE   2  (bases 1 to 3244)
  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: "Commun 
            Biol."; date: "2022-11-2"; volume: "5(1):1163. doi"; pages: " 
            10.1038/s42003-022-04152-8"
COMMENT     Sequence Label: U6-sgRNA-pcr4
FEATURES             Location/Qualifiers
     source          1..3244
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(425..1216,1217..1285))
                     /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     1048..1067
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1286..1390)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1458..1476
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1514..1536)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     1636..1655
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1849..1871
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1863..1880
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1864..1880
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        complement(2171..2419)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(2229..2248)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(2399..2419)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(2508..2524)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2508..2524)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2521..2543)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2532..2548
                     /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(2556..2562,2563..2580,2581..2586))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2601..2622
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2739..2756)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(2910..3244,1..254))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2990..3009)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
       61 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
      121 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
      181 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
      241 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
      301 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
      361 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
      421 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
      481 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
      541 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
      601 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
      661 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
      721 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
      781 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
      841 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
      901 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
      961 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     1021 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     1081 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     1141 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     1201 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     1261 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     1321 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     1381 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca
     1441 tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg
     1501 atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag
     1561 cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg
     1621 gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat atgcggtgtg
     1681 aaataccgca cagatgcgta aggagaaaat accgcatcag gcgccattcg ccattcaggc
     1741 tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga
     1801 aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac
     1861 gttgtaaaac gacggccagt gccaagcttg catgcctgca ggtcgacgat taaaaaaaag
     1921 ggctcgtccg ggatttgaac ccgggacctc tcgcacccta agcgagaatc atacccctag
     1981 accaacgagc cgcgatatac ctgctgctgg gaattgtagt atctacactt agtagaaatt
     2041 aaatctcacc ttttgagctg cttatctaca cttagtagaa attcattggc ctcaactgga
     2101 ctctacatct acacttagta gaaatttggg gctgaggctg ctgggttcca tctacactta
     2161 gtagaaattc ggtgtttcgt cctttccaca agatatataa agccaagaaa tcgaaatact
     2221 ttcaagttac ggtaagcata tgatagtcca ttttaaaaca taattttaaa actgcaaact
     2281 acccaagaaa ttattacttt ctacgtcacg tattttgtac taatatcttt gtgtttacag
     2341 tcaaattaat tctaattatc tctctaacag ccttgtatcg tatatgcaaa tatgaaggaa
     2401 tcatgggaaa taggccctct tcctgcccga ccttggtacc ggatccagtc gactgaattg
     2461 gaatctctag aggatccccg ggtaccgagc tcgaattcgt aatcatggtc atagctgttt
     2521 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     2581 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     2641 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     2701 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     2761 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     2821 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     2881 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     2941 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     3001 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     3061 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     3121 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     3181 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     3241 gact
//
