LOCUS       sequence-373799-        3011 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression, Genome editing.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3011)
  AUTHORS   Teng F, Cui T, Gao Q, Guo L, Zhou Q, Li W
  TITLE     Artificial sgRNAs engineered for genome editing with new Cas12b 
            orthologs.
  JOURNAL   Cell Discov. 2019 Apr 23;5:23. doi: 10.1038/s41421-019-0091-0. 
            eCollection 2019.
  PUBMED    31016029
REFERENCE   2  (bases 1 to 3011)
  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: "Cell 
            Discov."; date: "2019-04-23"; volume: "5:23. doi"; pages: " 
            10.1038/s41421-019-0091-0. eCollection 2019"
COMMENT     Sequence Label: pUC19-U6-Aa-artsg13 (BsaI)
FEATURES             Location/Qualifiers
     source          1..3011
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        402..650
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     402..422
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     573..592
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(790..806)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(790..806)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(803..825)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    814..830
                     /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(838..844,845..862,863..868))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    883..904
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1021..1038)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1192..1780)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1272..1291)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1951..2742,2743..2811))
                     /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     2574..2593
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2812..2916)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2984..3002
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt cgagggccta tttcccatga
      421 ttccttcata tttgcatata cgatacaagg ctgttagaga gataattgga attaatttga
      481 ctgtaaacac aaagatatta gtacaaaata cgtgacgtag aaagtaataa tttcttgggt
      541 agtttgcagt tttaaaatta tgttttaaaa tggactatca tatgcttacc gtaacttgaa
      601 agtatttcga tttcttggct ttatatatct tgtggaaagg acgaaacacc ggggtcgtct
      661 ataggacggc gagtttttca acgggtgtgc caatggccac tttccaggtg gcaaagcccg
      721 ttgagcttca aagaagtggc acagagaccg agagagggtc tcattttttt taagcttggc
      781 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
      841 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
      901 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
      961 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     1021 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     1081 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     1141 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     1201 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     1261 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     1321 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     1381 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     1441 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     1501 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     1561 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     1621 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     1681 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     1741 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     1801 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     1861 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     1921 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     1981 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     2041 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagagccacg
     2101 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     2161 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     2221 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     2281 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     2341 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     2401 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     2461 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     2521 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     2581 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     2641 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     2701 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     2761 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     2821 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     2881 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     2941 tgacgtctaa gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag
     3001 gccctttcgt c
//
