LOCUS       Exported                7032 bp ds-DNA     circular SYN 19-JAN-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7032)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Wednesday, Jul 5, 2023 from SnapGene 4.3.11
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7032
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        1..138
                     /label=Spn promoter
     CDS             139..1149
                     /label=SpnR
     rep_origin      1216..1889
                     /direction=RIGHT
                     /label=pUC origin
     terminator      2160..2187
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      2279..2365
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(2442..2460)
                     /label=pYPQ230-BB-Inv-R
     protein_bind    2461..2560
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     2461..2488
                     /label=Insert-Inv-F
     gene            2579..2629
                     /label=N-terminal NLS
                     /note="N-terminal NLS"
     CDS             2585..2605
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(2600..2629)
                     /label=pYPQ230-R
     CDS             2630..6403
                     /codon_start=1
                     /label=Ev1Cas12a (rice codon optimized)
                     /translation="ESNNKIFTETIGTSSIAKTMRNSLVPTESTKRNIEKNGIIIDDQL
                     RAEKRQQLKEIMDEYYRTYIDNKLSNVALTRTIDWKELFQAIEDNYKQNTTKTKNELEK
                     KQKEKRTEIYKILSDDEKFKQLFNAKLLTNVLPEFIKNQNIDNEEKQEKISTVELFQRF
                     TSSFTDFFKNRKNVFSKDEISTSICYRVVQENAWIFYQNLLAFEEIKKTAEQEIEKIEA
                     ENRDSISDYSLKEIFDFDFYGLLLNQGGIRFYNDVCGKINYHMNLYGQKHNIKSNKFKM
                     KRMHKQILSIDESTFEVPTMFENDKEVYQVLNEFLSDLASKKILERVEKIGENVSEYEI
                     NKIYIQSKNFEKFSSFMCGNWQIINDSLKTYYNEKIKSKGKAKEEKVKKAIKAIEYKSL
                     ADINQLVERYNNDELNRKAEEYISAINEKIKDLDVNEIEYDEKINLIENETKSEEIKSK
                     LDSIMEIMHWTKMFIIEEEIEKDVNFYNEIEEIYDELQPLVTIYNRIRNYVTQKPYSEE
                     KIKLNFGIPTLANGWSKTKEYDNNAIIMIRDGKYYLGIFNAKNKPDKKIMEGHQSEENG
                     DYKKMIYRLLPGPNKMLPKVFMSKTGIAEYKPSQYILECYEQNKHIKSDKNFDIKFCRD
                     LIDFFKTSINRHPEWSKFNFKFSETSEYEDISTFYREVEKQGYKIEWTYISEKEIKELD
                     ENGQLYLFQIYNKDFSEKSKGKENLHTMYLKNLFSEENLKNIVLKLNGEAEVFFRKSSI
                     KKPIIHKKGSVLVNKTYNENGERKSIPEEQYTEIYKYLNSIGTNELSEKSKKLMEEGKV
                     EYYKANYDIVKDYRYSVDKFFIHLPMTINFKAAGFSPINNIALKNIALKDDMHIIGIDR
                     GERNLIYVSVIDTKGNIVEQRNFNIVNGIDYKEKLKQKELDRDNARKNWKEIGKIKDLK
                     EGYLSLVVHEIAKLVVKYNAIITMEDLNQGFKRGRFKVERQVYQKFETMLINKLNYLVD
                     KDLAVDQEGGLLRGYQLTYIPESLKVLGRQCGYIFYVPAAYTSKIDPTTGFVAIFNYKG
                     MTDKDFVTSFDSIKYDDERGLFAFEFDYENFVTHKVEMARNKWTVYTYGERIKRKFKNG
                     SWDTAEKVDLTYQMRSILEKYEIEYNKGQDILEQIEELDEKAQNGICKEIKYLVKDIVQ
                     MRNSLPDNAAEDYDAIISPVINNNGEFFDSTRGDEDKPLDADANGAYCIALKGLYEVMQ
                     IKKNWNEETEFPRKELKIRHQDWFDFIQNKRYL"
     misc_feature    6404..6451
                     /label=Nucleoplasmin NLS
                     /note="Nucleoplasmin NLS"
     CDS             6404..6451
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     primer_bind     6404..6435
                     /label=pYPQ230-F
     misc_feature    6480..6757
                     /label=NOS term
     misc_feature    complement(6524..6547)
                     /label=Nos-term-R2
                     /note="Nos-term-R2"
     protein_bind    6779..6902
                     /gene="mutant version of attR"
                     /label=attR5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(6883..6902)
                     /label=Insert-Inv-R
     primer_bind     6903..6921
                     /label=pYPQ230-BB-Inv-F
     promoter        7031..7032
                     /label=Spn promoter
ORIGIN
        1 tggaccagcc aggacagaaa tgcctcgact tcgctgctac ccaaggttgc cgggtgacgc
       61 acaccgtgga aacggatgaa ggcacgaacc cagtggacat aagcctgttc ggttcgtaag
      121 ctgtaatgca agtagcgtat gcgctcacgc aactggtcca gaaccttgac cgaacgcagc
      181 ggtggtaacg gcgcagtggc ggttttcatg gcttgttatg actgtttttt tggggtacag
      241 tctatgcctc gggcatccaa gcagcaagcg cgttacgccg tgggtcgatg tttgatgtta
      301 tggagcagca acgatgttac gcagcagggc agtcgcccta aaacaaagtt aaacatcatg
      361 agggaagcgg tgatcgccga agtatcgact caactatcag aggtagttgg cgtcatcgag
      421 cgccatctcg aaccgacgtt gctggccgta catttgtacg gctccgcagt ggatggcggc
      481 ctgaagccac acagtgatat tgatttgctg gttacggtga ccgtaaggct tgatgaaaca
      541 acgcggcgag ctttgatcaa cgaccttttg gaaacttcgg cttcccctgg agagagcgag
      601 attctccgcg ctgtagaagt caccattgtt gtgcacgacg acatcattcc gtggcgttat
      661 ccagctaagc gcgaactgca atttggagaa tggcagcgca atgacattct tgcaggtatc
      721 ttcgagccag ccacgatcga cattgatctg gctatcttgc tgacaaaagc aagagaacat
      781 agcgttgcct tggtaggtcc agcggcggag gaactctttg atccggttcc tgaacaggat
      841 ctatttgagg cgctaaatga aaccttaacg ctatggaact cgccgcccga ctgggctggc
      901 gatgagcgaa atgtagtgct tacgttgtcc cgcatttggt acagcgcagt aaccggcaaa
      961 atcgcgccga aggatgtcgc tgccgactgg gcaatggagc gcctgccggc ccagtatcag
     1021 cccgtcatac ttgaagctag acaggcttat cttggacaag aagaagatcg cttggcctcg
     1081 cgcgcagatc agttggaaga atttgtccac tacgtgaaag gcgagatcac caaggtagtc
     1141 ggcaaataac cctcgagcca cccatgacca aaatccctta acgtgagtta cgcgtcgttc
     1201 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     1261 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     1321 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     1381 aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     1441 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     1501 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     1561 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     1621 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     1681 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     1741 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     1801 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     1861 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     1921 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
     1981 cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc
     2041 gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc
     2101 agtgagcgca acgcaattaa tacgcgtacc gcgagccagg aagagtttgt agaaacgcaa
     2161 aaaggccatc cgtcaggatg gccttctgct tagtttgatg cctggcagtt tatggcgggc
     2221 gtcctgcccg ccaccctccg ggccgttgct tcacaacgtt caaatccgct cccggcggat
     2281 ttgtcctact caggagagcg ttcaccgaca aacaacagat aaaacgaaag gcccagtctt
     2341 ccgactgagc ctttcgtttt atttgatgcc tggcagttcc ctactctcgc gttaacgctt
     2401 gcatggatgt tttcccagtc acgacgttgt aaaacgacgg ccagtcttaa gctcgggccc
     2461 caaataatga ttttattttg actgatagtg acctgttcgt tgcaacaaat tgatgagcaa
     2521 tgctttttta taatgccaac tttgtacaaa aaagcaggct ccgaattcgc ccttcaccat
     2581 ggctcctaag aagaagcgga aggttggtat tcacggggtg cctgcggctg aatcaaacaa
     2641 taaaatcttt accgaaacca tcggcacttc cagcatagcg aaaacgatgc gcaattcact
     2701 cgttcctacc gagagcacga agcgcaacat tgaaaaaaat gggattataa tagacgacca
     2761 actgagggcc gagaaacggc aacagctcaa agaaatcatg gacgaatact atcggactta
     2821 catagacaat aagctgtcga acgttgccct cacgagaaca atagattgga aggaactttt
     2881 tcaagcgatc gaagataatt acaagcaaaa cacgaccaag accaaaaacg aattggaaaa
     2941 aaagcaaaag gagaagcgga cggagattta caaaatcctc tctgacgacg aaaagttcaa
     3001 acagttgttc aatgcgaagc ttctcactaa cgttcttccg gaattcataa agaatcagaa
     3061 catagacaat gaggaaaaac aggaaaaaat ttctacagtc gaactttttc aaaggtttac
     3121 gagttctttc accgatttct ttaagaaccg gaagaacgtt ttctccaaag acgaaatctc
     3181 gacatcgatt tgctataggg ttgtccagga aaacgcatgg atattctatc agaatcttct
     3241 ggcttttgaa gagatcaaga aaacggctga acaggaaatc gagaagatcg aagcagagaa
     3301 ccgcgacagc atatctgact atagtttgaa ggaaattttc gatttcgatt tctacggcct
     3361 tctgctgaat cagggcggga tacgcttcta caatgacgtc tgcggcaaga ttaattatca
     3421 tatgaatttg tacggtcaaa aacacaatat aaagtcaaat aaatttaaaa tgaagcggat
     3481 gcataaacaa atattgtcga tagatgaatc gaccttcgaa gtgccgacga tgtttgagaa
     3541 cgataaagag gtgtaccagg tcctcaatga gtttctgtct gatctggctt ccaagaagat
     3601 cttggaaagg gtggaaaaaa taggggagaa cgtgagtgaa tacgagatca ataagattta
     3661 tattcaaagc aaaaattttg aaaaattctc ttcatttatg tgcgggaact ggcaaattat
     3721 caacgattca ctgaagactt attacaatga gaagataaag tcgaaaggta aagcaaagga
     3781 ggagaaagtc aaaaaagcta taaaggcaat cgaatataag tctcttgctg acataaacca
     3841 gcttgtcgag agatacaata acgacgaact gaatcgcaaa gcagaagagt atatatcggc
     3901 gataaatgag aaaatcaagg atctcgacgt gaatgaaatt gagtatgatg aaaaaattaa
     3961 tcttattgaa aatgagacta aatccgagga gatcaaaagc aagctcgaca gcataatgga
     4021 aatcatgcac tggacgaaaa tgtttatcat tgaagaggag atagaaaagg atgttaactt
     4081 ctataatgaa atagaggaga tttacgacga actgcaaccc cttgtcacga tctacaatcg
     4141 gatcaggaat tacgtcactc agaagcctta ttccgaagaa aagattaaac tgaacttcgg
     4201 aattccaaca cttgccaacg gctggagcaa aaccaaagaa tacgacaaca atgcgattat
     4261 catgatccgg gacggaaaat actacctcgg catcttcaat gccaagaata aaccggacaa
     4321 aaaaattatg gaaggacacc aaagcgagga gaatggcgat tacaaaaaga tgatctaccg
     4381 cttgcttcct ggccctaaca agatgctgcc aaaggttttc atgtctaaga ctggcatagc
     4441 ggagtataag ccgtctcagt acattctcga atgttatgag caaaataagc acataaaatc
     4501 tgacaaaaat tttgatataa aattttgtag agaccttatc gattttttca agacatcgat
     4561 taatcggcat cctgaatggt cgaagttcaa ctttaagttc tctgagacat ccgaatacga
     4621 agatatctca acattctacc gggaggttga aaaacaaggt tacaaaatag agtggaccta
     4681 catcagcgaa aaagaaatca aagaacttga cgaaaatggg cagttgtatt tgttccagat
     4741 ttataataag gatttttctg agaaaagtaa gggcaaggag aatctgcaca cgatgtattt
     4801 gaaaaatctg ttctcggaag agaatcttaa aaatatcgtc cttaaactta atggtgaagc
     4861 ggaggtgttt ttcaggaaat cttcaataaa aaagcccatt attcataaaa agggttccgt
     4921 tttggtcaac aagacgtata acgaaaacgg tgaacggaag tccataccag aggagcagta
     4981 tacagaaata tataaatatc ttaacagtat tggaactaat gagctgtcag aaaaatcgaa
     5041 gaaacttatg gaagaaggaa aagtcgaata ctataaagca aactacgaca ttgtgaaaga
     5101 ttaccgctat tcagtcgata aattctttat tcatctgcca atgacaataa attttaaggc
     5161 agccgggttt tcacctatta ataatattgc gttgaaaaac atcgccctga aagatgatat
     5221 gcatattata ggcatagatc ggggcgagcg gaatcttatc tatgtctccg tcattgatac
     5281 taaaggcaat atcgtcgaac agaggaattt taacatagtc aatggcattg attacaaaga
     5341 gaagttgaaa caaaaagagt tggatcggga caatgcgaga aaaaactgga aggagattgg
     5401 caagatcaag gaccttaaag agggctactt gtctctcgtc gttcatgaga ttgctaaact
     5461 ggttgtcaaa tataacgcaa taattactat ggaagacttg aatcaaggtt ttaaaagggg
     5521 cagatttaag gttgaacggc aagtgtacca gaagttcgaa acaatgttga taaataaatt
     5581 gaattatctg gtggacaagg acctcgccgt tgaccaggag ggagggctcc tgcggggcta
     5641 tcagttgacg tatatcccag agtctctcaa agtgctcgga cggcagtgcg ggtacatatt
     5701 ctacgtcccc gccgcataca cttctaaaat cgatccgaca acaggattcg tggctatatt
     5761 taattataaa ggaatgacag acaaagactt cgtcacatcg tttgactcaa taaaatatga
     5821 cgacgaacgg ggcctctttg ctttcgaatt cgactatgag aatttcgtca cgcacaaggt
     5881 cgaaatggca cggaataagt ggacggtcta cacatacgga gaaaggataa agagaaagtt
     5941 caaaaatggc agctgggata ccgcagagaa agttgacctg acataccaga tgagatctat
     6001 actggagaag tatgaaatag agtacaacaa ggggcaagac atcttggaac aaatagaaga
     6061 actcgatgag aaggcgcaaa acggcatatg caaggagatt aaatatctgg tcaaggacat
     6121 cgtccaaatg cgcaactccc tcccagacaa cgctgccgag gattatgatg caatcattag
     6181 tcctgtgatt aacaacaatg gggagttctt cgactcaaca aggggggacg aggacaaacc
     6241 gcttgatgca gacgcgaatg gcgcatattg tattgctttg aagggactgt acgaggttat
     6301 gcaaataaaa aaaaattgga atgaagagac ggaatttccg cgcaaggaac tcaagatacg
     6361 gcatcaggat tggttcgatt ttatccaaaa caaaagatac ctgaagcgtc ctgctgccac
     6421 caaaaaggcc ggacaggcta agaaaaagaa gtgagacgac tagtggcggc cgccgacgtc
     6481 cgatcgttca aacatttggc aataaagttt cttaagattg aatcctgttg ccggtcttgc
     6541 gatgattatc atataatttc tgttgaatta cgttaagcat gtaataatta acatgtaatg
     6601 catgacgtta tttatgagat gggtttttat gattagagtc ccgcaattat acatttaata
     6661 cgcgatagaa aacaaaatat agcgcgcaaa ctaggataaa ttatcgcgcg cggtgtcatc
     6721 tatgttacta gatcgggaat tgatcccccc tcgacagctt ccggaaaggg cgaattcgca
     6781 actttgtata caaaagttga acgagaaacg taaaatgata taaatatcaa tatattaaat
     6841 tagattttgc ataaaaaaca gactacataa tactgtaaaa cacaacatat ccagtcacta
     6901 tgccatccag ctgatatccc ctatagtgag tcgtattaca tggtcatagc tgtttcctgg
     6961 cagctctggc ccgtgtctca aaatctctga tgttacattg cacaagataa aaatatatca
     7021 tcatgcctcc tc
//
