LOCUS       sequence-235875-        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     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(857..873)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(857..873)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(870..892)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    881..897
                     /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(905..911,912..929,930..935))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    950..971
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1088..1105)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1259..1847)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1339..1358)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2018..2809,2810..2878))
                     /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     2641..2660
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2879..2983)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3051..3069
                     /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 ggaacaaagc
      661 accagtggtc tagtggtgga atagtaccct gccacggtac agacccgggt tcgattcccg
      721 gctggtgcaa gagaccgaga gagggtctca aacaaagcac cagtggtcta gtggtggaat
      781 agtaccctgc cacggtacag acccgggttc gattcccggc tggtgcattt ttttttttaa
      841 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
      901 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
      961 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     1021 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
     1081 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     1141 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     1201 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     1261 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     1321 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     1381 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     1441 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     1501 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     1561 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     1621 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     1681 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     1741 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     1801 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     1861 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     1921 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     1981 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     2041 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     2101 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgtg
     2161 agccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     2221 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     2281 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     2341 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     2401 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     2461 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     2521 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     2581 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     2641 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     2701 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     2761 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     2821 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     2881 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     2941 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     3001 tgccacctga cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta
     3061 tcacgaggcc ctttcgtc
//
