LOCUS       sequence-411203-        7255 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses NG cas9C by the constitutive CASI promoter and sgRNA 
            targeting murine Lmna c.1621T mutation by U6 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7255)
  AUTHORS   Yang L, Liu Z, Sun J, Chen Z, Gao F, Guo Y
  TITLE     Adenine base editor-based correction of the cardiac pathogenic Lmna 
            c.1621C > T mutation in murine hearts.
  JOURNAL   J Cell Mol Med. 2024 Feb;28(4):e18145. doi: 10.1111/jcmm.18145.
  PUBMED    38332517
REFERENCE   2  (bases 1 to 7255)
  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: "J Cell Mol 
            Med."; date: "2024-02"; volume: "28(4):e18145. doi"; pages: " 
            10.1111/jcmm.18145"
COMMENT     Sequence Label: pAAV-CASI-InteinC-NG C aa714-1368-U6-Lmna sgRNA1
FEATURES             Location/Qualifiers
     source          1..7255
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   12..141
                     /label=AAV2 ITR
     enhancer        217..520
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        522..799
                     /label=chicken beta-actin promoter
     regulatory      1273..1282
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      1277..1286
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3442..3462
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    3477..3532
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3531..3550)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     misc_RNA        complement(3637..3712)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(3733..3981)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(3791..3810)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(3961..3981)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     repeat_region   4316..4445
                     /label=AAV2 ITR
     primer_bind     complement(4464..4481)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4464..4480)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4473..4495)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4622..5077
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4709..4728)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4919..4940
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5103..5207
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5208..5276,5277..6068)
                     /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     complement(5426..5445)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6239..6827
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6728..6747
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    7115..7136
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7151..7156,7157..7174,7175..7181)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7189..7205
                     /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)."
     primer_bind     7194..7216
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7213..7229
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7213..7229
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ccttaattag gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc ttgtagttaa tgattaaccc gccatgctac ttatctacgt
      181 agccatgctc taggaagatc ggaattcgga gttccgcgtt acataactta cggtaaatgg
      241 cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc
      301 catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac
      361 tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa
      421 tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac
      481 ttggcagtac atctacgtat tagtcatcgc tattaccatg gtcgaggtga gccccacgtt
      541 ctgcttcact ctccccatct cccccccctc cccaccccca attttgtatt tatttatttt
      601 ttaattattt tgtgcagcga tgggggcggg gggggggggg gggsgcgcgc sagggggggg
      661 gggggggggg gagggggggg gsggggsgag gcggagaggt gcggcggcag ccaatcagag
      721 cggcgcgctc cgaaagtttc cttttatggc gaggcggcgg cggcggcggc cctataaaaa
      781 gcgaagcgcg cggcgggcgg gagtcgctgc gcgctgcctt cgccccgtgc cccgctccgc
      841 cgccgcctcg cgccgcccgc cccggctctg actgaccgcg ttactaaaac aggtaagtcc
      901 ggcctccgcg ccgggttttg gcgcctcccg cgggcgcccc cctcctcacg gcgagcgctg
      961 ccacgtcaga cgaagggcgc agcgagcgtc ctgatccttc cgcccggacg ctcaggacag
     1021 cggcccgctg ctcataagac tcggccttag aaccccagta tcagcagaag gacattttag
     1081 gacgggactt gggtgactct agggcactgg ttttctttcc agagagcgga acaggcgagg
     1141 aaaagtagtc ccttctcggc gattctgcgg agggatctcc gtggggcggt gaacgccgat
     1201 gatgcctcta ctaaccatgt tcatgttttc tttttttttc tacaggtcct gggtgacgaa
     1261 caggctagcg ccgccaccat ggcggcggcg tgcccggaac tgcgtcagct ggcgcagagc
     1321 gatgtgtatt gggatccgat tgtgagcatt gaaccggatg gcgtggaaga agtgtttgat
     1381 ctgaccgtgc cgggcccgca taactttgtg gcgaacgata ttattgcgca taactctggc
     1441 cagggcgata gcctgcacga gcacattgcc aatctggccg gcagccccgc cattaagaag
     1501 ggcatcctgc agacagtgaa ggtggtggac gagctcgtga aagtgatggg ccggcacaag
     1561 cccgagaaca tcgtgatcga aatggccaga gagaaccaga ccacccagaa gggacagaag
     1621 aacagccgcg agagaatgaa gcggatcgaa gagggcatca aagagctggg cagccagatc
     1681 ctgaaagaac accccgtgga aaacacccag ctgcagaacg agaagctgta cctgtactac
     1741 ctgcagaatg ggcgggatat gtacgtggac caggaactgg acatcaaccg gctgtccgac
     1801 tacgatgtgg accatatcgt gcctcagagc tttctgaagg acgactccat cgacaacaag
     1861 gtgctgacca gaagcgacaa gaaccggggc aagagcgaca acgtgccctc cgaagaggtc
     1921 gtgaagaaga tgaagaacta ctggcggcag ctgctgaacg ccaagctgat tacccagaga
     1981 aagttcgaca atctgaccaa ggccgagaga ggcggcctga gcgaactgga taaggccggc
     2041 ttcatcaaga gacagctggt ggaaacccgg cagatcacaa agcacgtggc acagatcctg
     2101 gactcccgga tgaacactaa gtacgacgag aatgacaagc tgatccggga agtgaaagtg
     2161 atcaccctga agtccaagct ggtgtccgat ttccggaagg atttccagtt ttacaaagtg
     2221 cgcgagatca acaactacca ccacgcccac gacgcctacc tgaacgccgt cgtgggaacc
     2281 gccctgatca aaaagtaccc taagctggaa agcgagttcg tgtacggcga ctacaaggtg
     2341 tacgacgtgc ggaagatgat cgccaagagc gagcaggaaa tcggcaaggc taccgccaag
     2401 tacttcttct acagcaacat catgaacttt ttcaagaccg agattaccct ggccaacggc
     2461 gagatccgga agcggcctct gatcgagaca aacggcgaaa ccggggagat cgtgtgggat
     2521 aagggccggg attttgccac cgtgcggaaa gtgctgagca tgccccaagt gaatatcgtg
     2581 aaaaagaccg aggtgcagac aggcggcttc agcaaagagt ctatcaggcc caagaggaac
     2641 agcgataagc tgatcgccag aaagaaggac tgggacccta agaagtacgg cggcttcgtc
     2701 agccccaccg tggcctattc tgtgctggtg gtggccaaag tggaaaaggg caagtccaag
     2761 aaactgaaga gtgtgaaaga gctgctgggg atcaccatca tggaaagaag cagcttcgag
     2821 aagaatccca tcgactttct ggaagccaag ggctacaaag aagtgaaaaa ggacctgatc
     2881 atcaagctgc ctaagtactc cctgttcgag ctggaaaacg gccggaagag aatgctggcc
     2941 tctgccagat tcctgcagaa gggaaacgaa ctggccctgc cctccaaata tgtgaacttc
     3001 ctgtacctgg ccagccacta tgagaagctg aagggctccc ccgaggataa tgagcagaaa
     3061 cagctgtttg tggaacagca caagcactac ctggacgaga tcatcgagca gatcagcgag
     3121 ttctccaaga gagtgatcct ggccgacgct aatctggaca aagtgctgtc cgcctacaac
     3181 aagcaccggg ataagcccat cagagagcag gccgagaata tcatccacct gtttaccctg
     3241 accaatctgg gagcccctag ggccttcaag tactttgaca ccaccatcga ccggaaggtg
     3301 tacaggagca ccaaagaggt gctggacgcc accctgatcc accagagcat caccggcctg
     3361 tacgagacac ggatcgacct gtctcagctg ggaggtgact ctggcggctc aaaaagaacc
     3421 gccgacggca gcgaattcga gcccaagaag aagaggaaag tctaaaccgg ttggctaata
     3481 aaggaaattt attttcattg caatagtgtg ttggaatttt ttgtgtctct cactcggaag
     3541 gacatatggg agggcaaatc atttaaaaca tcagaatgag tatttggttt agagtttggc
     3601 aacatatgcc catatgaggc cctcacgcgt aaaaaagcac cgactcggtg ccactttttc
     3661 aagttgataa cggactagcc ttattttaac ttgctatttc tagctctaaa acggaagtgg
     3721 ccatgtgcaa gcggtgtttc gtcctttcca caagatatat aaagccaaga aatcgaaata
     3781 ctttcaagtt acggtaagca tatgatagtc cattttaaaa cataatttta aaactgcaaa
     3841 ctacccaaga aattattact ttctacgtca cgtattttgt actaatatct ttgtgtttac
     3901 agtcaaatta attccaatta tctctctaac agccttgtat cgtatatgca aatatgaagg
     3961 aatcatggga aataggccct cggtaccggc gatgctggct gccatgaaca aaggttggct
     4021 ataaagaggt catcagtata tgaaacagcc ccctgctgtc cattccttat tccatagaaa
     4081 agccttgact tgaggttaga ttttttttat attttgtttt gtgttatttt tttctttaac
     4141 atccctaaaa ttttccttac atgttttact agccagattt ttcctcctct cctgactact
     4201 cccagtcata gctgtccctc ttctcttatg gagatcggat ccgaattccc gataaggatc
     4261 ttcctagagc atggctacgt agataagtag catggcgggt taatcattaa ctacaaggaa
     4321 cccctagtga tggagttggc cactccctct ctgcgcgctc gctcgctcac tgaggccggg
     4381 cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg
     4441 cgcagcctta attaacctaa ttcactggcc gtcgttttac aacgtcgtga ctgggaaaac
     4501 cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat
     4561 agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg
     4621 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     4681 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     4741 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     4801 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     4861 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     4921 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     4981 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     5041 aacgcgaatt ttaacaaaat attaacgctt acaatttagg tggcactttt cggggaaatg
     5101 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     5161 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac
     5221 atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc
     5281 cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca
     5341 tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc
     5401 caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg
     5461 ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac
     5521 cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca
     5581 taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg
     5641 agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac
     5701 cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg
     5761 caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat
     5821 taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg
     5881 ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg
     5941 cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc
     6001 aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc
     6061 attggtaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt
     6121 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     6181 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     6241 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     6301 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     6361 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
     6421 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     6481 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     6541 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     6601 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     6661 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     6721 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     6781 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     6841 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
     6901 tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc
     6961 gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcccaatac
     7021 gcaaaccgcc tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc
     7081 ccgactggaa agcgggcagt gagcgcaacg caattaatgt gagttagctc actcattagg
     7141 caccccaggc tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat
     7201 aacaatttca cacaggaaac agctatgacc atgattacgc cagatttaat taagg
//
