LOCUS       Exported                8899 bp ds-DNA     circular SYN 18-JAN-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8899)
  AUTHORS   locusliu
  TITLE     Direct Submission
  JOURNAL   Exported Sunday, Apr 4, 2021 from SnapGene Viewer 5.2.4
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8899
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     regulatory      626..635
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             638..664
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             704..724
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     misc_feature    749..3904
                     /label=SaCas9-KKH-N580A
     primer_bind     1274..1296
                     /label=kkh-seq1
     primer_bind     1908..1929
                     /label=kkh-seq2
     primer_bind     2531..2554
                     /label=kkh-seq3
     primer_bind     3088..3110
                     /label=kkh-seq4
     primer_bind     3704..3726
                     /label=kkh-seq5
     misc_feature    4004..6070
                     /label=M-MLV
     misc_feature    6071..6124
                     /label=BPSV40
     CDS             6134..6154
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6163..6180
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    6209..6433
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6504..6520)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6528..6544
                     /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(6552..6582)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6597..6618
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(6906..7494)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(7665..8525)
                     /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"
     promoter        complement(8526..8630)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc
      601 agatccgcta gagatccgcg gccgcgccac catggggccc gccgccaagc gcgtgaagct
      661 ggacggcggc aaacggacag ccgacggaag cgagttcgag tcaccaaaga agaagcggaa
      721 agtcggtatc cacggagtcc cagcagccaa acggaactac atcctggggc ttgacattgg
      781 gataaccagc gttggctacg gaattattga ttatgagaca cgcgatgtga ttgacgccgg
      841 ggttaggctg ttcaaagagg ccaacgttga aaacaacgag ggaagacgga gtaagcgcgg
      901 agcaagaaga ctcaagcgca gacggagaca tcggattcag agggtgaaaa agctgctctt
      961 cgattacaat ctcctgaccg atcatagtga gctgagcgga atcaacccct acgaggcgcg
     1021 agtgaaaggg ctttcccaga agctgtccga agaggagttc tccgccgcgt tgctgcacct
     1081 ggccaaacgg aggggggttc acaatgtaaa cgaagtggag gaggacacgg gcaatgaact
     1141 tagtacgaaa gaacagatca gtaggaactc taaggctctc gaagagaaat acgtcgctga
     1201 gttgcagctt gagagactga aaaaagacgg cgaagtacgc ggatctatta ataggttcaa
     1261 gacttcagat tacgtaaagg aagccaagca gctcctgaaa gtacagaaag cgtaccatca
     1321 gctcgatcag agcttcatcg atacctacat agatttgctg gagacacgga ggacatacta
     1381 cgagggccca ggggaaggat ctccttttgg gtggaaggac atcaaggaat ggtacgagat
     1441 gcttatggga cattgtacat attttccgga ggagctcagg agcgtcaagt acgcctacaa
     1501 tgccgacctg tacaatgccc tcaatgacct caataacctc gtgattacca gggacgagaa
     1561 cgagaagctg gagtactatg aaaagttcca gattatcgag aatgtgttta agcagaagaa
     1621 gaagccgaca cttaagcaga ttgcaaagga aatcctcgtg aatgaggaag atatcaaggg
     1681 atacagagtg acaagtacag gcaagcccga gttcacaaat ctgaaggtgt accacgatat
     1741 taaggacata accgcacgaa aggagataat cgaaaacgct gagctcctcg atcagatcgc
     1801 aaaaattctt accatctacc agtctagtga ggacattcag gaggaactga ctaatctgaa
     1861 cagtgagctc acccaagagg aaattgagca gatttcaaac ctgaaaggct acaccgggac
     1921 gcacaatctg agcctcaaag caatcaacct cattctggat gaactttggc acacaaatga
     1981 caaccaaatt gccatattca accgcctgaa actggtgcca aaaaaagtgg atctgtcaca
     2041 gcaaaaggaa atccctacaa ccttggttga cgattttatt ctgtcccccg ttgtcaagcg
     2101 gagcttcatc cagtcaatca aggtgatcaa tgccatcatt aaaaaatacg gattgccaaa
     2161 cgatataatt atcgagcttg cacgagagaa gaactcaaag gacgcccaga agatgattaa
     2221 cgaaatgcag aagcgcaacc gccagacaaa cgaacgcata gaggaaatta taagaacaac
     2281 cggcaaagag aatgccaagt atctgatcga gaaaatcaag ctgcacgaca tgcaagaagg
     2341 caagtgcctg tactctctgg aagctatccc actcgaagat ctgctgaata atccattcaa
     2401 ttacgaggtg gaccacatca tccctagatc cgtaagcttt gacaattcct tcaataacaa
     2461 agttctggtt aaacaggagg aagcctctaa aaaagggaac cggaccccgt tccagtacct
     2521 gagctccagt gacagcaaga ttagctacga gacttttaag aaacatattc tgaatctggc
     2581 caaaggcaaa ggcaggatca gcaagaccaa gaaggagtac ctcctcgaag aacgcgacat
     2641 taacagattt agtgtgcaga aagatttcat caaccgaaac cttgtcgata ctcggtacgc
     2701 cacgagaggc ctgatgaatc tcctcaggag ctacttccgc gtcaataatc tggacgttaa
     2761 agtcaagagc ataaatgggg gattcaccag ctttctgagg agaaagtgga agtttaagaa
     2821 ggaacgaaac aaaggataca agcaccatgc tgaggatgct ttgatcatcg ctaacgcgga
     2881 ctttatcttt aaggaatgga aaaagctgga taaggcaaag aaagtgatgg aaaaccagat
     2941 gttcgaggag aagcaggcag agtcaatgcc tgagatcgag acagagcagg aatacaagga
     3001 aattttcatc acccctcatc agattaaaca cataaaggac ttcaaagact ataaatactc
     3061 tcatagggtg gacaaaaaac ccaatcgcaa gctcattaat gacaccctgt actcaacacg
     3121 gaaggatgat aaaggtaata ccttgattgt gaataatctt aatggattgt atgacaaaga
     3181 taacgacaag ctcaagaagc tgatcaacaa gtctccagag aagctcctta tgtatcacca
     3241 cgacccacag acttatcaga aattgaaact gatcatggag caatacgggg atgagaagaa
     3301 cccactctac aaatattatg aggaaacagg taattacctg accaagtact ccaagaagga
     3361 taacggacca gtgatcaaaa agataaagta ctatggcaac aaacttaatg cgcatttgga
     3421 cataactgac gattacccca attctcgaaa caaggttgtg aagctctccc tgaagcctta
     3481 tagatttgac gtgtacctgg ataatggggt ttataaattc gtcaccgtga aaaatctgga
     3541 cgtgatcaaa aaggagaact attatgaagt aaactcaaag tgctatgagg aggcgaagaa
     3601 gctgaagaag atctccaatc aggccgagtt catcgcttcc ttctataaga acgatctcat
     3661 caagatcaat ggagagcttt atcgcgtcat tggtgtgaac aatgacttgc tgaacaggat
     3721 cgaagtcaat atgatagaca ttacctaccg ggagtatctc gaaaacatga atgataaacg
     3781 gccgcctcac atcatcaaga caatcgcatc taaaactcag tcaataaaaa agtactctac
     3841 cgatatcctg gggaatctct atgaagtgaa gtcaaagaag cacccacaaa tcattaaaaa
     3901 aggttctgga ggatctagcg gaggatcctc tggcagcgag acaccaggaa caagcgagtc
     3961 agcaacacca gagagcagtg gcggcagcag cggcggcagc agcaccctaa atatagaaga
     4021 tgagtatcgg ctacatgaga cctcaaaaga gccagatgtt tctctagggt ccacatggct
     4081 gtctgatttt cctcaggcct gggcggaaac cgggggcatg ggactggcag ttcgccaagc
     4141 tcctctgatc atacctctga aagcaacctc tacccccgtg tccataaaac aataccccat
     4201 gtcacaagaa gccagactgg ggatcaagcc ccacatacag agactgttgg accagggaat
     4261 actggtaccc tgccagtccc cctggaacac gcccctgcta cccgttaaga aaccagggac
     4321 taatgattat aggcctgtcc aggatctgag agaagtcaac aagcgggtgg aagacatcca
     4381 ccccaccgtg cccaaccctt acaacctctt gagcgggctc ccaccgtccc accagtggta
     4441 cactgtgctt gatttaaagg atgccttttt ctgcctgaga ctccacccca ccagtcagcc
     4501 tctcttcgcc tttgagtgga gagatccaga gatgggaatc tcaggacaat tgacctggac
     4561 cagactccca cagggtttca aaaacagtcc caccctgttt aatgaggcac tgcacagaga
     4621 cctagcagac ttccggatcc agcacccaga cttgatcctg ctacagtacg tggatgactt
     4681 actgctggcc gccacttctg agctagactg ccaacaaggt actcgggccc tgttacaaac
     4741 cctagggaac ctcgggtatc gggcctcggc caagaaagcc caaatttgcc agaaacaggt
     4801 caagtatctg gggtatcttc taaaagaggg tcagagatgg ctgactgagg ccagaaaaga
     4861 gactgtgatg gggcagccta ctccgaagac ccctcgacaa ctaagggagt tcctagggaa
     4921 ggcaggcttc tgtcgcctct tcatccctgg gtttgcagaa atggcagccc ccctgtaccc
     4981 tctcaccaaa ccggggactc tgtttaattg gggcccagac caacaaaagg cctatcaaga
     5041 aatcaagcaa gctcttctaa ctgccccagc cctggggttg ccagatttga ctaagccctt
     5101 tgaactcttt gtcgacgaga agcagggcta cgccaaaggt gtcctaacgc aaaaactggg
     5161 accttggcgt cggccggtgg cctacctgtc caaaaagcta gacccagtag cagctgggtg
     5221 gcccccttgc ctacggatgg tagcagccat tgccgtactg acaaaggatg caggcaagct
     5281 aaccatggga cagccactag tcattctggc cccccatgca gtagaggcac tagtcaaaca
     5341 accccccgac cgctggcttt ccaacgcccg gatgactcac tatcaggcct tgcttttgga
     5401 cacggaccgg gtccagttcg gaccggtggt agccctgaac ccggctacgc tgctcccact
     5461 gcctgaggaa gggctgcaac acaactgcct tgatatcctg gccgaagccc acggaacccg
     5521 acccgaccta acggaccagc cgctcccaga cgccgaccac acctggtaca cggatggaag
     5581 cagtctctta caagagggac agcgtaaggc gggagctgcg gtgaccaccg agaccgaggt
     5641 aatctgggct aaagccctgc cagccgggac atccgctcag cgggctgaac tgatagcact
     5701 cacccaggcc ctaaagatgg cagaaggtaa gaagctaaat gtttatactg atagccgtta
     5761 tgcttttgct actgcccata tccatggaga aatatacaga aggcgtgggt ggctcacatc
     5821 agaaggcaaa gagatcaaaa ataaagacga gatcttggcc ctactaaaag ccctctttct
     5881 gcccaaaaga cttagcataa tccattgtcc aggacatcaa aagggacaca gcgccgaggc
     5941 tagaggcaac cggatggctg accaagcggc ccgaaaggca gccatcacag agactccaga
     6001 cacctctacc ctcctcatag aaaattcatc accctctggc ggctcaaaaa gaaccgccga
     6061 cggcagcgag aagcgcaccg ccgacagcca gcacagcacc ccccccaaga ccaagcgcaa
     6121 ggtggaattc gagcccaaga agaagaggaa agtctaaccg gtcatcatca ccatcaccat
     6181 tgagtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt
     6241 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     6301 taaaatgaga aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     6361 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg
     6421 gtgggctcta tggcttctga ggcggaaaga accagctggg gctcgatacc gtcgacctct
     6481 agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     6541 acaattccac acaacatacg agccggaagc ataaagtgta aagcctaggg tgcctaatga
     6601 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     6661 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     6721 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     6781 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     6841 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     6901 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     6961 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     7021 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     7081 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     7141 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     7201 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     7261 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     7321 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     7381 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     7441 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     7501 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     7561 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     7621 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     7681 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     7741 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     7801 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     7861 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     7921 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     7981 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     8041 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     8101 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     8161 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     8221 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     8281 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     8341 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     8401 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     8461 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     8521 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     8581 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     8641 cgaaaagtgc cacctgacgt cgacggatcg ggagatcgat ctcccgatcc cctagggtcg
     8701 actctcagta caatctgctc tgatgccgca tagttaagcc agtatctgct ccctgcttgt
     8761 gtgttggagg tcgctgagta gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg
     8821 accgacaatt gcatgaagaa tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta
     8881 cgggccagat atacgcgtt
//
