LOCUS       sequence-390162-        3226 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  nicking sgRNA to induce LMNA (c.1824C > T) mutation using PE3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3226)
  AUTHORS   Li M, Zhong A, Wu Y, Sidharta M, Beaury M, Zhao X, Studer L, Zhou T
  TITLE     Transient inhibition of p53 enhances prime editing and cytosine 
            base-editing efficiencies in human pluripotent stem cells.
  JOURNAL   Nat Commun. 2022 Oct 27;13(1):6354. doi: 10.1038/s41467-022-34045-7.
  PUBMED    36302757
REFERENCE   2  (bases 1 to 3226)
  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: "2022-10-27"; volume: "13(1):6354. doi"; pages: " 
            10.1038/s41467-022-34045-7"
COMMENT     Sequence Label: Lsg-LMNA-1 + 2
FEATURES             Location/Qualifiers
     source          1..3226
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     7..26
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        7..25
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        40..288
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     40..60
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     misc_RNA        310..385
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(419..439)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(419..437)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(458..474)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(458..474)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(471..493)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    482..498
                     /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(506..512,513..530,531..536))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    551..572
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(689..706)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(860..1448)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(940..959)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1619..2410,2411..2479))
                     /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     2242..2261
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2480..2584)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2610..3065)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2747..2768)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2959..2978
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3192..3214
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3206..3223
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3207..3223
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 cgcgcgtaat acgactcact atagggcgaa ttgggtaccg agggcctatt tcccatgatt
       61 ccttcatatt tgcatatacg atacaaggct gttagagaga taattggaat taatttgact
      121 gtaaacacaa agatattagt acaaaatacg tgacgtagaa agtaataatt tcttgggtag
      181 tttgcagttt taaaattatg ttttaaaatg gactatcata tgcttatcgt aacttgaaag
      241 tatttcgatt tcttggcttt atatatcttg tggaaaggac gaaacaccgc cccactgccc
      301 cccacactgg ttttagagct agaaatagca agttaaaata aggctagtcc gttatcaact
      361 tgaaaaagtg gcaccgagtc ggtgcttttt ttccgcggtg gagctccagc ttttgttccc
      421 tttagtgagg gttaattgcg cgcttggcgt aatcatggtc atagctgttt cctgtgtgaa
      481 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
      541 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
      601 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
      661 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
      721 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
      781 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
      841 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
      901 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
      961 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     1021 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     1081 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     1141 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     1201 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     1261 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     1321 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     1381 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     1441 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     1501 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     1561 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     1621 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     1681 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     1741 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     1801 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     1861 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     1921 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     1981 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     2041 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     2101 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     2161 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     2221 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     2281 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     2341 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     2401 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     2461 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     2521 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     2581 cgcgcacatt tccccgaaaa gtgccaccta aattgtaagc gttaatattt tgttaaaatt
     2641 cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat
     2701 cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa
     2761 gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg
     2821 cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa
     2881 agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc
     2941 gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag
     3001 tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg
     3061 cgcgtcccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc
     3121 ttcgctatta cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac
     3181 gccagggttt tcccagtcac gacgttgtaa aacgacggcc agtgag
//
