LOCUS       sequence-322133-        3413 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  plasmid expressing an sgRNA targeting the BEAR-GFP plasmid along 
            with an mCherry marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3413)
  AUTHORS   Talas A, Simon DA, Kulcsar PI, Varga E, Krausz SL, Welker E
  TITLE     BEAR reveals that increased fidelity variants can successfully 
            reduce the mismatch tolerance of adenine but not cytosine base 
            editors.
  JOURNAL   Nat Commun. 2021 Nov 3;12(1):6353. doi: 10.1038/s41467-021-26461-y.
  PUBMED    34732717
REFERENCE   2  (bases 1 to 3413)
  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: "Nat 
            Commun."; date: "2021-11-3"; volume: "12(1):6353. doi"; pages: " 
            10.1038/s41467-021-26461-y"
COMMENT     Sequence Label: pAT15415-BEAR-GFP-target-mCherry
FEATURES             Location/Qualifiers
     source          1..3413
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        53..301
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     53..73
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     224..243
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        322..397
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        432..643
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      663..672
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             669..1379
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(814..832)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     complement(1409..1426)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1415..1622
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(1687..2275)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1767..1786)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2446..3237,3238..3306))
                     /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     3069..3088
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3307..3411)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 catttccccg aaaagtgcca cctgacgtcc agcagagatc cactttggcg ccgagggcct
       61 atttcccatg attccttcat atttgcatat acgatacaag gctgttagag agataattgg
      121 aattaatttg actgtaaaca caaagatatt agtacaaaat acgtgacgta gaaagtaata
      181 atttcttggg tagtttgcag ttttaaaatt atgttttaaa atggactatc atatgcttac
      241 cgtaacttga aagtatttcg atttcttggc tttatatatc ttgtggaaag gacgaaacac
      301 cgcagacaag tgcatagact ggttttagag ctagaaatag caagttaaaa taaggctagt
      361 ccgttatcaa cttgaaaaag tggcaccgag tcggtgcttt tttggctcga gtggctccgg
      421 tgcccgtcag tgggcagagc gcacatcgcc cacagtcccc gagaagttgg ggggaggggt
      481 cggcaattga accggtgcct agagaaggtg gcgcggggta aactgggaaa gtgatgtcgt
      541 gtactggctc cgcctttttc ccgagggtgg gggagaaccg tatataagtg cagtagtcgc
      601 cgtgaacgtt ctttttcgca acgggtttgc cgccagaaca caggtgtcgt gacgcgggat
      661 ccgccaccat ggtgagcaag ggcgaggagg ataacatggc catcatcaag gagttcatgc
      721 gcttcaaggt gcacatggag ggctccgtga acggccacga gttcgagatc gagggcgagg
      781 gcgagggccg cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggtggcc
      841 ccctgccctt cgcctgggac atcctgtccc ctcagttcat gtacggctcc aaggcctacg
      901 tgaagcaccc cgccgacatc cccgactact tgaagctgtc cttccccgag ggcttcaagt
      961 gggagcgcgt gatgaacttc gaggacggcg gcgtggtgac cgtgacccag gactcctccc
     1021 tgcaggacgg cgagttcatc tacaaggtga agctgcgcgg caccaacttc ccctccgacg
     1081 gccccgtaat gcagaaaaag accatgggct gggaggcctc ctccgagcgg atgtaccccg
     1141 aggacggcgc cctgaagggc gagatcaagc agaggctgaa gctgaaggac ggcggccact
     1201 acgacgctga ggtcaagacc acctacaagg ccaagaagcc cgtgcagctg cccggcgcct
     1261 acaacgtcaa catcaagttg gacatcacct cccacaacga ggactacacc atcgtggaac
     1321 agtacgaacg cgccgagggc cgccactcca ccggcggcat ggacgagctg tacaagtaga
     1381 tctacatgtc ctagagctcg ctgatcagcc tcgactgtgc cttctagttg ccagccatct
     1441 gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt
     1501 tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg
     1561 ggtggggtgg ggcaggacag caagggggag gattgggaag agaatagcag gcatgctggg
     1621 gagcggccgc tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     1681 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     1741 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     1801 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     1861 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     1921 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     1981 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     2041 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     2101 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     2161 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     2221 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     2281 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     2341 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     2401 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     2461 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     2521 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     2581 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     2641 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     2701 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     2761 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     2821 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     2881 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     2941 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     3001 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     3061 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     3121 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     3181 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     3241 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     3301 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     3361 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gca
//
