LOCUS       sequence-350382-        4613 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  BEAR-GFP plasmid with a gRNA that targets its own plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4613)
  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 4613)
  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: pAT15516_BEAR-GFP-2in1
FEATURES             Location/Qualifiers
     source          1..4613
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        52..300
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     52..72
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     223..242
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        321..396
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        457..760
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        761..964
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     915..935
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1003..1012
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(1054..1075)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2106..2125)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2453..2474
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    2517..2582
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    2706..2827
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2743..2762)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2797..2816
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2886..3474)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2966..2985)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3645..4436,4437..4505))
                     /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     4268..4287
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4506..4610)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 atttccccga aaagtgccac ctgacgtcca gcagagatcc actttggcgc cgagggccta
       61 tttcccatga ttccttcata tttgcatata cgatacaagg ctgttagaga gataattgga
      121 attaatttga ctgtaaacac aaagatatta gtacaaaata cgtgacgtag aaagtaataa
      181 tttcttgggt agtttgcagt tttaaaatta tgttttaaaa tggactatca tatgcttacc
      241 gtaacttgaa agtatttcga tttcttggct ttatatatct tgtggaaagg acgaaacacc
      301 gcagacaagt gcatagactg gttttagagc tagaaatagc aagttaaaat aaggctagtc
      361 cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt ttggctcgag tggctccggt
      421 gcccgtcagt gggcagagcg cacatcgccc acagtccgtt acataactta cggtaaatgg
      481 cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc
      541 catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac
      601 tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa
      661 tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac
      721 ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta
      781 catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga
      841 cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa
      901 ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag
      961 agctggttta gtgaaccgtc agatccgcta gcgctaccgg tcgccaccat ggtgagcaag
     1021 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
     1081 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
     1141 ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
     1201 ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc
     1261 ttcaagtccg ccatgcccga aggctacgtg cagacaagtg catagactgc gggttgattt
     1321 tttagttaaa atatgggaaa gtttaaaaga aggaatgatg aataaactat acaaccactt
     1381 taaatatgtg ttggtcccct ttaagtcagc aagcagatat atcagatatg gcagctaact
     1441 taaggagtcc ccgtggatac tggatgaata gcccccaatt ccaccaatgc aagaaccaac
     1501 agtctatttg gtcttgatta ggacagtaaa cttcctgcac accatagtct gtgcactaaa
     1561 caggagtaaa acacccttaa ggcatctctt acattggctg cattaaagtg tgtcacttaa
     1621 tgacatccgt gatatcaggg atagtttgac attggacata cagttctctc tcagtgattg
     1681 tcacctgctt atatagtacg aggacccata tgtggtcaat cactttcttg ttgccactcc
     1741 atgccaggga agacatagtt acaatgagtt ataattataa aagctcagcc ttgcatgagg
     1801 atataaaata catttgccat acctctacat atgccttagt tgcttttaag ctttgtttgg
     1861 aattgcaaag gccaacttcc taaacacagt ttcaaagtat tcttttgtca cacctttcca
     1921 ctggatactt gtattgtctg caggaagagg ccaccctgcc tatgcttctc cttagggcct
     1981 cgttctgcag agtcagcata aactcccagt ctgctcaacc aattctgtct gattagggcc
     2041 atatctcctt accatgagtt gatttttgtt tcttttttga gcaggagcgc accatcttct
     2101 tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg
     2161 tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca
     2221 agctggagta caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg
     2281 gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg
     2341 accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact
     2401 acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc
     2461 tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagtccg
     2521 gactcagatc tcgagctcaa gcttcgaatt ctgcagtcga cggtaccgcg ggcccgggat
     2581 ccaccggatc tagataactg atcataatca gccataccac atttgtagag gttttacttg
     2641 ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg
     2701 ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     2761 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     2821 tatcttagct gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     2881 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     2941 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     3001 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     3061 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     3121 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     3181 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     3241 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     3301 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     3361 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     3421 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     3481 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     3541 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     3601 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     3661 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     3721 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     3781 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     3841 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     3901 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     3961 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     4021 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     4081 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     4141 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     4201 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     4261 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     4321 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     4381 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     4441 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     4501 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     4561 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cac
//
