LOCUS       sequence-240582-        3078 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression, Genome editing, backbone.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3078)
  AUTHORS   Teng F, Li J, Cui T, Xu K, Guo L, Gao Q, Feng G, Chen C, Han D, Zhou
            Q, Li W
  TITLE     Enhanced mammalian genome editing by new Cas12a orthologs with 
            optimized crRNA scaffolds.
  JOURNAL   Genome Biol. 2019 Feb 5;20(1):15. doi: 10.1186/s13059-019-1620-8.
  PUBMED    30717767
REFERENCE   2  (bases 1 to 3078)
  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: "Genome 
            Biol."; date: "2019-02-5"; volume: "20(1):15. doi"; pages: " 
            10.1186/s13059-019-1620-8"
COMMENT     Sequence Label: pUC19-U6-tRNAx2
FEATURES             Location/Qualifiers
     source          1..3078
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     56..78
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     70..87
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     71..87
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        94..342
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     94..114
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     265..284
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(549..565)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(549..565)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(562..584)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    573..589
                     /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(597..603,604..621,622..627))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    642..663
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(780..797)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(951..1539)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1031..1050)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1710..2501,2502..2570))
                     /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     2333..2352
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2571..2675)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2743..2761
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2799..2821)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2921..2940
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag
       61 tcacgacgtt gtaaaacgac ggccagtgaa ttcgagggcc tatttcccat gattccttca
      121 tatttgcata tacgatacaa ggctgttaga gagataattg gaattaattt gactgtaaac
      181 acaaagatat tagtacaaaa tacgtgacgt agaaagtaat aatttcttgg gtagtttgca
      241 gttttaaaat tatgttttaa aatggactat catatgctta ccgtaacttg aaagtatttc
      301 gatttcttgg ctttatatat cttgtggaaa ggacgaaaca ccggaacaaa gcaccagtgg
      361 tctagtggtg gaatagtacc ctgccacggt acagacccgg gttcgattcc cggctggtgc
      421 aagagaccga gagagggtct caaacaaagc accagtggtc tagtggtgga atagtaccct
      481 gccacggtac agacccgggt tcgattcccg gctggtgcat tttttttttt aagcttggcg
      541 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
      601 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
      661 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
      721 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
      781 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
      841 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
      901 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
      961 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     1021 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     1081 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     1141 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     1201 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     1261 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     1321 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     1381 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     1441 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     1501 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     1561 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     1621 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     1681 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     1741 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     1801 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agagccacgc
     1861 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     1921 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     1981 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     2041 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     2101 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     2161 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     2221 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     2281 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     2341 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     2401 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     2461 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     2521 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     2581 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     2641 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     2701 gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg
     2761 ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg
     2821 gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg
     2881 tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta
     2941 ctgagagtgc accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc
     3001 atcaggcgcc attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc
     3061 tcttcgctat tacgccag
//
