LOCUS       Exported                7149 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 7149)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Wednesday, Jul 5, 2023 from SnapGene 4.3.11
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7149
                     /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..6520
                     /codon_start=1
                     /label=Hs1Cas12a (rice codon optimized)
                     /translation="TKTFDSEFFNLYSLQKTVRFELKPVGETASFVEDFKNEGLKRVVS
                     EDERRAVDYQKVKEIIDDYHRDFIEESLNYFPEQVSKDALEQAFHLYQKLKAAKVEERE
                     KALKEWEALQKKLREKVVKCFSDSNKARFSRIDKKELIKEDLINWLVAQNREDDIPTVE
                     TFNNFTTYFTGFHENRKNIYSKDDHATAISFRLIHENLPKFFDNVISFNKLKEGFPELK
                     FDKVKEDLEVDYDLKHAFEIEYFVNFVTQAGIDQYNYLLGGKTLEDGTKKQGMNEQINL
                     FKQQQTRDKARQIPKLIPLFKQILSERTESQSFIPKQFESDQELFDSLQKLHNNCQDKF
                     TVLQQAILGLAEADLKKVFIKTSDLNALSNTIFGNYSVFSDALNLYKESLKTKKAQEAF
                     EKLPAHSIHDLIQYLEQFNSSLDAEKQQSTDTVLNYFIKTDELYSRFIKSTSEAFTQVQ
                     PLFELEALSSKRRPPESEDEGAKGQEGFEQIKRIKAYLDTLMEAVHFAKPLYLVKGRKM
                     IEGLDKDQSFYEAFEMAYQELESLIIPIYNKARSYLSRKPFKADKFKINFDNNTLLSGW
                     DANKETANASILFKKDGLYYLGIMPKGKTFLFDYFVSSEDSEKLKQRRQKTAEEALAQD
                     GESYFEKIRYKLLPGASKMLPKVFFSNKNIGFYNPSDDILRIRNTASHTKNGTPQKGHS
                     KVEFNLNDCHKMIDFFKSSIQKHPEWGSFGFTFSDTSDFEDMSAFYREVENQGYVISFD
                     KIKETYIQSQVEQGNLYLFQIYNKDFSPYSKGKPNLHTLYWKALFEEANLNNVVAKLNG
                     EAEIFFRRHSIKASDKVVHPANQAIDNKNPHTEKTQSTFEYDLVKDKRYTQDKFFFHVP
                     ISLNFKAQGVSKFNDKVNGFLKGNPDVNIIGIDRGERHLLYFTVVNQKGEILVQESLNT
                     LMSDKGHVNDYQQKLDKKEQERDAARKSWTTVENIKELKEGYLSHVVHKLAHLIIKYNA
                     IVCLEDLNFGFKRGRFKVEKQVYQKFEKALIDKLNYLVFKEKELGEVGHYLTAYQLTAP
                     FESFKKLGKQSGILFYVPADYTSKIDPTTGFVNFLDLRYQSVEKAKQLLSDFNAIRFNS
                     VQNYFEFEIDYKKLTPKRKVGTQSKWVICTYGDVRYQNRRNQKGHWETEEVNVTEKLKA
                     LFASDSKTTTVIDYANDDNLIDVILEQDKASFFKELLWLLKLTMTLRHSKIKSEDDFIL
                     SPVKNEQGEFYDSRKAGEVWPKDADANGAYHIALKGLWNLQQINQWEKGKTLNLAIKNQ
                     DWFSFIQEKPYQE"
     CDS             2911..2922
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             6521..6568
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     misc_feature    6521..6568
                     /label=Nucleoplasmin NLS
                     /note="Nucleoplasmin NLS"
     primer_bind     6521..6552
                     /label=pYPQ230-F
     misc_feature    6597..6874
                     /label=NOS term
     misc_feature    complement(6641..6664)
                     /label=Nos-term-R2
                     /note="Nos-term-R2"
     protein_bind    6896..7019
                     /gene="mutant version of attR"
                     /label=attR5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(7000..7019)
                     /label=Insert-Inv-R
     primer_bind     7020..7038
                     /label=pYPQ230-BB-Inv-F
     promoter        7148..7149
                     /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 cctgcggcta ccaagacatt
     2641 cgactctgaa tttttcaacc tttacagtct tcagaaaacc gtcagatttg agctcaaacc
     2701 ggtcggtgaa acggcctcat tcgtcgagga ctttaagaac gaaggcctga agcgggttgt
     2761 tagcgaagac gagaggcgcg cagtggatta tcaaaaagtg aaggaaatca ttgacgatta
     2821 tcatcgcgat ttcattgaag agtcactgaa ctacttccca gaacaggttt caaaagacgc
     2881 cttggaacag gcttttcact tgtaccaaaa attgaaggcc gccaaggtgg aagaacggga
     2941 gaaagccttg aaggagtggg aagcgctcca aaaaaaattg agggagaaag tggtgaaatg
     3001 ctttagtgat tcaaacaagg ccaggtttag tagaatagat aaaaaggaac tcataaagga
     3061 ggatcttatt aattggctcg tggctcaaaa ccgcgaagac gacattccca cagtggaaac
     3121 cttcaacaac ttcaccactt atttcaccgg gtttcacgaa aatagaaaga atatatacag
     3181 caaggatgac catgcaaccg ctatatcttt ccgcttgata cacgaaaacc tcccgaaatt
     3241 cttcgacaat gtcatctcgt tcaacaagct caaggaaggt tttcccgaat tgaaattcga
     3301 taaggtgaaa gaagacttgg aggtggacta tgacctgaaa catgcctttg aaatagaata
     3361 ttttgtcaat ttcgtgacac aagcggggat cgatcaatat aattacttgc tgggcggcaa
     3421 gacgttggag gatggcacta aaaagcaggg catgaatgag cagatcaatc ttttcaagca
     3481 acagcaaact agagacaagg ctagacagat cccaaaattg atcccattgt tcaaacagat
     3541 cctttcggag aggaccgaat ctcagtcgtt tatccccaaa cagttcgaaa gtgaccagga
     3601 actgtttgat tcccttcaaa aattgcataa caactgtcag gacaagttca ctgtcttgca
     3661 gcaagccatc ttgggcctgg cggaagcaga tttgaaaaaa gtgtttataa agacctcgga
     3721 cttgaatgcg cttagcaata cgatttttgg caattactct gtgtttagcg atgcactcaa
     3781 cctttataaa gagagcctga agaccaagaa agcccaagaa gcttttgaga aacttcctgc
     3841 gcattcgatc catgatctta ttcaatatct tgagcagttt aactcaagtc ttgacgcgga
     3901 aaagcagcag tccactgaca cggttctgaa ctatttcatc aaaacagatg aactctactc
     3961 gagatttatt aaatcaacct cggaggcatt tactcaggtg caaccgcttt tcgagctcga
     4021 agccctttcc agcaaacgca gaccaccgga atccgaagat gaaggtgcta aaggacaaga
     4081 agggttcgaa caaattaagc ggattaaggc atatcttgat acgcttatgg aagcggtcca
     4141 cttcgcgaag ccgctttacc tggtcaaggg gcggaaaatg atcgagggtc ttgacaaaga
     4201 tcagtctttt tacgaggcgt ttgagatggc ttaccaggag ctcgaaagtt tgataatccc
     4261 aatatacaac aaggccagat cctatctctc gcggaagcct tttaaggcag ataagttcaa
     4321 gattaacttt gacaacaaca cgcttttgtc aggctgggac gccaataagg aaactgccaa
     4381 cgcgagtatc ctctttaaaa aagacggtct ttactatctt ggtataatgc caaaaggtaa
     4441 aacatttttg tttgattatt tcgtcagctc ggaagattcc gaaaaactta aacagcggag
     4501 gcaaaagacg gccgaggagg ccctcgcgca ggatggtgag agctattttg aaaaaattcg
     4561 ctataagttg ctccccggag ctagtaaaat gttgcccaag gtcttcttta gtaataagaa
     4621 cattggcttc tacaatccct cagatgatat ccttaggata aggaatacag cgagccacac
     4681 gaaaaacggc acaccgcaaa aaggacattc caaagtggag ttcaatctga acgattgcca
     4741 taagatgatc gatttcttta aatcctctat ccaaaaacac cccgaatggg gctcgttcgg
     4801 tttcactttt tcggacacga gtgattttga agacatgagt gcgttctata gggaggttga
     4861 aaatcaaggg tatgtcatct cgtttgataa gattaaagaa acgtatattc aaagccaggt
     4921 tgaacagggc aatctctatc ttttccagat atacaataaa gacttttcgc catactctaa
     4981 gggaaagccg aatctccata ctctgtattg gaaagccctc ttcgaggaag cgaaccttaa
     5041 caatgttgtg gcaaagctta atggggaagc agaaatattt ttccgccgcc actcaataaa
     5101 ggccagcgac aaagttgtgc atcctgcgaa ccaggccatt gataataaga acccgcacac
     5161 agaaaagact cagagtacct tcgaatatga tcttgttaaa gacaagcgct acacacagga
     5221 caagttcttt ttccacgttc caatatcact caatttcaag gcgcaaggag tcagcaaatt
     5281 taatgataaa gtcaatggat tcctcaaagg gaatccggat gtgaacatta ttgggataga
     5341 cagaggcgag agacacctgc tgtactttac ggttgtgaat caaaaggggg aaattttggt
     5401 ccaggagtct ctcaacacgc tcatgagtga caagggccac gtcaatgatt accaacagaa
     5461 gctggacaaa aaagaacagg agagggatgc agcaagaaaa agctggacga cggtggagaa
     5521 tatcaaggag ctgaaagaag gttacttgtc gcacgttgtg cataagctgg ctcatctgat
     5581 cattaaatac aatgccatcg tctgcttgga ggacctcaac tttggtttta aacgcgggag
     5641 atttaaggtc gagaaacagg tttaccaaaa gttcgaaaaa gcgctcatcg ataagctgaa
     5701 ctacctggtg tttaaagaaa aagaacttgg ggaagtcggg cactatctca cagcgtacca
     5761 gcttacagcg ccttttgaga gcttcaaaaa actcggcaaa cagtcgggca tccttttcta
     5821 cgtcccggcg gattatacgt caaagataga ccccacaaca gggtttgtta acttcttgga
     5881 tcttcggtat caaagcgtcg aaaaggctaa gcaacttctc tccgacttca acgctataag
     5941 gtttaatagc gtccaaaatt atttcgagtt tgaaatagat tataagaagc tcacaccaaa
     6001 gcggaaggtt gggacccagt ctaagtgggt catatgcaca tacggcgatg tgaggtacca
     6061 gaataggcgc aatcaaaagg gtcactggga gaccgaagag gttaatgtga ctgagaagct
     6121 taaggctctt tttgcctcag actctaaaac aactactgtg atcgattacg ccaacgatga
     6181 caacctcatt gacgtgatcc tcgaacaaga taaggcctcc ttttttaaag aactcctttg
     6241 gcttctgaag ctgacgatga cactccgcca ctccaaaatc aagagcgagg acgactttat
     6301 actgtcacct gtgaagaatg aacaaggcga attttacgac tcgaggaaag caggcgaagt
     6361 ttggcccaaa gacgctgacg ccaatggggc atatcacatc gctcttaagg gactctggaa
     6421 cttgcaacag attaatcagt gggagaaagg gaagaccttg aatctcgcga taaagaatca
     6481 agattggttc agctttattc aagaaaagcc gtaccaagaa aagcgtcctg ctgccaccaa
     6541 aaaggccgga caggctaaga aaaagaagtg agacgactag tggcggccgc cgacgtccga
     6601 tcgttcaaac atttggcaat aaagtttctt aagattgaat cctgttgccg gtcttgcgat
     6661 gattatcata taatttctgt tgaattacgt taagcatgta ataattaaca tgtaatgcat
     6721 gacgttattt atgagatggg tttttatgat tagagtcccg caattataca tttaatacgc
     6781 gatagaaaac aaaatatagc gcgcaaacta ggataaatta tcgcgcgcgg tgtcatctat
     6841 gttactagat cgggaattga tcccccctcg acagcttccg gaaagggcga attcgcaact
     6901 ttgtatacaa aagttgaacg agaaacgtaa aatgatataa atatcaatat attaaattag
     6961 attttgcata aaaaacagac tacataatac tgtaaaacac aacatatcca gtcactatgc
     7021 catccagctg atatccccta tagtgagtcg tattacatgg tcatagctgt ttcctggcag
     7081 ctctggcccg tgtctcaaaa tctctgatgt tacattgcac aagataaaaa tatatcatca
     7141 tgcctcctc
//
