LOCUS       Exported                6719 bp ds-DNA    circular SYN 13-9-2021
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    TS-PE2-N
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6719)
  AUTHORS   hyewon
  TITLE     Direct Submission
  JOURNAL   Exported 2021¤¤¤Å¤¤ 9 13¤·¤Ó¤© ¤·¤Í¤©¤·¤Ë¤·¤Ó¤© from SnapGene 2.3.2
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6719
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     misc_feature    1..130
                     /note="ITR"
     misc_feature    39..130
                     /note="AAV2 ITR"
     enhancer        154..533
                     /note="CMV enhancer"
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        534..737
                     /note="CMV promoter"
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     misc_feature    756..764
                     /note="kozak"
     CDS             798..818
                     /codon_start=1
                     /locus_tag="
                     "
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /note="SV40 NLS"
                     /note="
                     "
                     /protein_id="
                     "
                     /translation="PKKKRKV"
     CDS             819..3923
                     /codon_start=1
                     /locus_tag="
                     "
                     /product="nickase mutant of the Cas9 endonuclease from the 
                     Streptococcus pyogenes Type II CRISPR/Cas system"
                     /note="Cas9(H840A)"
                     /note="generates RNA-guided single strand nicks in DNA due 
                     to the H840A mutation in the HNH catalytic domain"
                     /protein_id="
                     "
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKY"
     misc_feature    3333..3335
                     /note="H840A"
     misc_feature    3924..3973
                     /note="SD"
     repeat_region   3982..4122
                     /note="AAV2 ITR"
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     rep_origin      4197..4652
                     /direction=RIGHT
                     /note="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        4934..5038
                     /gene="bla"
                     /note="AmpR promoter"
     CDS             5039..5899
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /note="AmpR"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      6070..6658
                     /direction=RIGHT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggcctcta gactcgaggc gttgacattg attattgact agttattaat
      181 agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac
      241 ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa
      301 tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt
      361 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc
      421 ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat
      481 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc
      541 ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc
      601 tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa
      661 aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg
      721 tctatataag cagagctctc tggctaacta ccggtgccac catgaaacgg acagccgacg
      781 gaagcgagtt cgagtcacca aagaagaagc ggaaagtcga caagaagtac agcatcggcc
      841 tggacatcgg caccaactct gtgggctggg ccgtgatcac cgacgagtac aaggtgccca
      901 gcaagaaatt caaggtgctg ggcaacaccg accggcacag catcaagaag aacctgatcg
      961 gagccctgct gttcgacagc ggcgaaacag ccgaggccac ccggctgaag agaaccgcca
     1021 gaagaagata caccagacgg aagaaccgga tctgctatct gcaagagatc ttcagcaacg
     1081 agatggccaa ggtggacgac agcttcttcc acagactgga agagtccttc ctggtggaag
     1141 aggataagaa gcacgagcgg caccccatct tcggcaacat cgtggacgag gtggcctacc
     1201 acgagaagta ccccaccatc taccacctga gaaagaaact ggtggacagc accgacaagg
     1261 ccgacctgcg gctgatctat ctggccctgg cccacatgat caagttccgg ggccacttcc
     1321 tgatcgaggg cgacctgaac cccgacaaca gcgacgtgga caagctgttc atccagctgg
     1381 tgcagaccta caaccagctg ttcgaggaaa accccatcaa cgccagcggc gtggacgcca
     1441 aggccatcct gtctgccaga ctgagcaaga gcagacggct ggaaaatctg atcgcccagc
     1501 tgcccggcga gaagaagaat ggcctgttcg gaaacctgat tgccctgagc ctgggcctga
     1561 cccccaactt caagagcaac ttcgacctgg ccgaggatgc caaactgcag ctgagcaagg
     1621 acacctacga cgacgacctg gacaacctgc tggcccagat cggcgaccag tacgccgacc
     1681 tgtttctggc cgccaagaac ctgtccgacg ccatcctgct gagcgacatc ctgagagtga
     1741 acaccgagat caccaaggcc cccctgagcg cctctatgat caagagatac gacgagcacc
     1801 accaggacct gaccctgctg aaagctctcg tgcggcagca gctgcctgag aagtacaaag
     1861 agattttctt cgaccagagc aagaacggct acgccggcta cattgacggc ggagccagcc
     1921 aggaagagtt ctacaagttc atcaagccca tcctggaaaa gatggacggc accgaggaac
     1981 tgctcgtgaa gctgaacaga gaggacctgc tgcggaagca gcggaccttc gacaacggca
     2041 gcatccccca ccagatccac ctgggagagc tgcacgccat tctgcggcgg caggaagatt
     2101 tttacccatt cctgaaggac aaccgggaaa agatcgagaa gatcctgacc ttccgcatcc
     2161 cctactacgt gggccctctg gccaggggaa acagcagatt cgcctggatg accagaaaga
     2221 gcgaggaaac catcaccccc tggaacttcg aggaagtggt ggacaagggc gcttccgccc
     2281 agagcttcat cgagcggatg accaacttcg ataagaacct gcccaacgag aaggtgctgc
     2341 ccaagcacag cctgctgtac gagtacttca ccgtgtataa cgagctgacc aaagtgaaat
     2401 acgtgaccga gggaatgaga aagcccgcct tcctgagcgg cgagcagaaa aaggccatcg
     2461 tggacctgct gttcaagacc aaccggaaag tgaccgtgaa gcagctgaaa gaggactact
     2521 tcaagaaaat cgagtgcttc gactccgtgg aaatctccgg cgtggaagat cggttcaacg
     2581 cctccctggg cacataccac gatctgctga aaattatcaa ggacaaggac ttcctggaca
     2641 atgaggaaaa cgaggacatt ctggaagata tcgtgctgac cctgacactg tttgaggaca
     2701 gagagatgat cgaggaacgg ctgaaaacct atgcccacct gttcgacgac aaagtgatga
     2761 agcagctgaa gcggcggaga tacaccggct ggggcaggct gagccggaag ctgatcaacg
     2821 gcatccggga caagcagtcc ggcaagacaa tcctggattt cctgaagtcc gacggcttcg
     2881 ccaacagaaa cttcatgcag ctgatccacg acgacagcct gacctttaaa gaggacatcc
     2941 agaaagccca ggtgtccggc cagggcgata gcctgcacga gcacattgcc aatctggccg
     3001 gcagccccgc cattaagaag ggcatcctgc agacagtgaa ggtggtggac gagctcgtga
     3061 aagtgatggg ccggcacaag cccgagaaca tcgtgatcga aatggccaga gagaaccaga
     3121 ccacccagaa gggacagaag aacagccgcg agagaatgaa gcggatcgaa gagggcatca
     3181 aagagctggg cagccagatc ctgaaagaac accccgtgga aaacacccag ctgcagaacg
     3241 agaagctgta cctgtactac ctgcagaatg ggcgggatat gtacgtggac caggaactgg
     3301 acatcaaccg gctgtccgac tacgatgtgg acgctatcgt gcctcagagc tttctgaagg
     3361 acgactccat cgacaacaag gtgctgacca gaagcgacaa gaaccggggc aagagcgaca
     3421 acgtgccctc cgaagaggtc gtgaagaaga tgaagaacta ctggcggcag ctgctgaacg
     3481 ccaagctgat tacccagaga aagttcgaca atctgaccaa ggccgagaga ggcggcctga
     3541 gcgaactgga taaggccggc ttcatcaaga gacagctggt ggaaacccgg cagatcacaa
     3601 agcacgtggc acagatcctg gactcccgga tgaacactaa gtacgacgag aatgacaagc
     3661 tgatccggga agtgaaagtg atcaccctga agtccaagct ggtgtccgat ttccggaagg
     3721 atttccagtt ttacaaagtg cgcgagatca acaactacca ccacgcccac gacgcctacc
     3781 tgaacgccgt cgtgggaacc gccctgatca aaaagtaccc taagctggaa agcgagttcg
     3841 tgtacggcga ctacaaggtg tacgacgtgc ggaagatgat cgccaagagc gagcaggaaa
     3901 tcggcaaggc taccgccaag tacgtaagta tcaaggttac aagacaggtt taaggagacc
     3961 aatagaaact ggggcggccg caggaacccc tagtgatgga gttggccact ccctctctgc
     4021 gcgctcgctc gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc
     4081 gggcggcctc agtgagcgag cgagcgcgca gctgcctgca ggggcgcctg atgcggtatt
     4141 ttctccttac gcatctgtgc ggtatttcac accgcatacg tcaaagcaac catagtacgc
     4201 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     4261 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     4321 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     4381 tttacggcac ctcgacccca aaaaacttga tttgggtgat ggttcacgta gtgggccatc
     4441 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     4501 cttgttccaa actggaacaa cactcaaccc tatctcgggc tattcttttg atttataagg
     4561 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     4621 gaattttaac aaaatattaa cgtttacaat tttatggtgc actctcagta caatctgctc
     4681 tgatgccgca tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg
     4741 ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat
     4801 gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg
     4861 cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt
     4921 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     4981 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat
     5041 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt
     5101 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg
     5161 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga
     5221 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg
     5281 tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt
     5341 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg
     5401 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg
     5461 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga
     5521 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc
     5581 tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc
     5641 ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc
     5701 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtggaagccg
     5761 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac
     5821 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc
     5881 actgattaag cattggtaac tgtcagacca agtttactca tatatacttt agattgattt
     5941 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
     6001 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     6061 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     6121 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     6181 aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg
     6241 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     6301 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     6361 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     6421 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     6481 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     6541 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     6601 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     6661 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgt
//
