LOCUS       sequence-272069-        7374 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway entry plasmid (attL1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7374)
  AUTHORS   Zhang Y, Ren Q, Tang X, Liu S, Malzahn AA, Zhou J, Wang J, Yin D, 
            Pan C, Yuan M, Huang L, Yang H, Zhao Y, Fang Q, Zheng X, Tian L, 
            Cheng Y, Le Y, McCoy B, Franklin L, Selengut JD, Mount SM, Que Q, 
            Zhang Y, Qi Y
  TITLE     Expanding the scope of plant genome engineering with Cas12a 
            orthologs and highly multiplexable editing systems.
  JOURNAL   Nat Commun. 2021 Mar 29;12(1):1944. doi: 10.1038/s41467-021-22330-w.
  PUBMED    33782402
REFERENCE   2  (bases 1 to 7374)
  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: "Nat 
            Commun."; date: "2021-03-29"; volume: "12(1):1944. doi"; pages: " 
            10.1038/s41467-021-22330-w"
COMMENT     Sequence Label: pYPQ281 (MbCas12a)
FEATURES             Location/Qualifiers
     source          1..7374
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             358..1149
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      1242..1830
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1731..1750
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1984..2001
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(2160..2187)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(2279..2365)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     2414..2436
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2428..2445
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2429..2445
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     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"
     CDS             2585..2605
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6746..6793
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     terminator      6824..7076
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     protein_bind    7121..7244
                     /gene="mutant version of attR"
                     /label=attR5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(7261..7280)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(7262..7280)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(7285..7301)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7285..7301)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
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 aaacattatg
      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 aatactgttc 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 cctgcggctc tgttccaaga
     2641 tttcactcat ctgtaccccc tctccaaaac agtgcggttt gaattgaagc cgatcgatcg
     2701 gacacttgaa cacattcacg caaaaaactt cctctctcag gatgagacca tggcagacat
     2761 gcaccagaaa gtcaaagtca tactcgatga ctaccatagg gacttcatag ccgacatgat
     2821 gggtgaggtt aaactgacta aactcgcgga attttacgat gtgtacttga aattcagaaa
     2881 aaatcccaag gacgacgagt tgcaaaagca actcaaggat ctccaggccg tcttgcgcaa
     2941 agaaattgtt aagccgatag gtaacggtgg caaatataag gcaggatacg acagactttt
     3001 cggtgctaaa ttgttcaaag acggtaaaga gctgggcgat ttggcaaagt ttgtgatagc
     3061 gcaagaaggt gagtcgtctc ccaaactggc ccatcttgcc cacttcgaga aattttcaac
     3121 gtactttaca ggatttcatg ataacagaaa aaacatgtac agcgacgagg ataaacacac
     3181 cgctattgct tataggttga tccatgaaaa tctgccgagg ttcatagata acctgcagat
     3241 ccttacgact attaaacaga aacatagtgc actttatgat caaataatca acgaactgac
     3301 tgcgtccggg ctggacgtgt cacttgcgtc acatctggat ggctatcaca agttgttgac
     3361 ccaggagggt atcaccgctt acaatacatt gcttggtggt atttcagggg aggctgggtc
     3421 gccaaaaatt caggggatca acgagctcat aaatagccat cacaatcagc actgccacaa
     3481 atcagaaagg attgccaaac tgcgccctct ccacaaacaa atcttgtcag atggaatgtc
     3541 ggtctccttt cttccgagca agtttgcaga cgactcggag atgtgccaag ctgtgaacga
     3601 attttacaga cattatgccg atgttttcgc caaggtccaa tcactctttg atggattcga
     3661 tgatcatcag aaggatggga tctatgttga acacaaaaac ctcaatgagc tctcgaagca
     3721 ggcgttcggg gatttcgctc tcttgggaag ggttttggat gggtattatg ttgatgtggt
     3781 gaatccggaa ttcaacgagc gcttcgctaa agcgaagacg gacaacgcga aagctaaact
     3841 cacaaaggag aaggacaagt tcatcaaggg tgtccactcg ttggccagcc tggaacaagc
     3901 aatagaacac tataccgcac gccacgatga tgagagtgtg caagcaggga agcttgggca
     3961 atactttaag catgggctcg cgggtgtgga caatcctatc cagaagatac acaacaacca
     4021 cagtacgatc aaagggtttc tggaaagaga acgccccgcg ggagagcggg cgcttccgaa
     4081 aattaagagc ggtaaaaatc ccgaaatgac ccagcttcgc cagttgaaag aacttctcga
     4141 taatgctttg aacgttgccc actttgctaa acttctgacc acgaaaacga cactcgataa
     4201 tcaagacggc aatttctacg gagagtttgg tgttctttat gacgaacttg cgaaaatccc
     4261 tacactctac aataaagttc gggattacct gtcccaaaaa ccattctcga cggagaaata
     4321 caagctgaat tttggcaatc ccactcttct gaacggatgg gacctgaaca aagaaaagga
     4381 taatttcgga gtgattctcc agaaggacgg ctgctactat cttgccctcc tggataaagc
     4441 tcataaaaaa gtgttcgata atgccccgaa tactggtaaa tctatttacc aaaaaatgat
     4501 atataaatat cttgaggttc ggaagcagtt cccgaaggtt ttttttagta aggaagcaat
     4561 cgcgataaac taccacccgt cgaaagagct cgtcgaaatt aaagataagg gcaggcaaag
     4621 gagtgacgac gaacggctta agctctatcg gtttatcttg gagtgtctca aaatccaccc
     4681 gaagtacgac aagaaatttg aaggcgctat tggtgacata cagctcttca aaaaggataa
     4741 aaagggaaga gaagtgccaa tatcagagaa agatttgttc gataagataa atggtatatt
     4801 tagtagtaag ccgaaattgg agatggaaga cttcttcatc ggggagttca agagatataa
     4861 tcccagccaa gatttggttg atcaatacaa catttataaa aagatagact ctaatgataa
     4921 cagaaagaaa gaaaatttct ataataatca ccctaaattc aagaaagatc tcgtccgcta
     4981 ctattatgag tctatgtgca agcacgagga atgggaggag tcttttgagt tctccaagaa
     5041 gctccaagac ataggttgtt atgtcgacgt gaacgaactt tttacggaga tcgagacccg
     5101 ccgcctgaac tataagatct cattttgcaa tatcaatgcg gattatatag acgagcttgt
     5161 ggaacaaggc cagttgtatc tttttcaaat ctacaacaaa gacttctcgc ctaaggctca
     5221 tggcaagccg aatctccaca cgttgtattt taaggcactt ttctctgagg acaacttggc
     5281 ggatccgata tacaagctca acggcgaggc ccagatcttt taccgcaaag cttcactcga
     5341 tatgaatgaa acaaccattc acagagctgg tgaggtgctc gagaacaaaa accccgacaa
     5401 tcccaagaaa aggcagtttg tgtatgacat aataaaagac aagcgctata cccaagacaa
     5461 atttatgctt cacgttccga tcacaatgaa cttcggtgtg cagggtatga ccattaagga
     5521 gttcaataaa aaagtgaatc agtccatcca gcaatatgat gaagtcaatg ttatcggaat
     5581 cgatcgggga gaacgccacc tgctctacct cacggttatt aatagcaaag gagaaatttt
     5641 ggagcaatgc tcgctgaatg acattacgac agcctccgct aacggaactc agatgactac
     5701 tccatatcat aagatactcg acaaaaggga aatagagaga ttgaatgcaa gggttggatg
     5761 gggtgaaatt gaaaccatta aggagctcaa gtctggttat ctgtctcacg ttgttcacca
     5821 gatttcacag ttgatgttga agtataacgc aattgtcgtg ctggaggatc ttaacttcgg
     5881 ctttaaaagg ggcaggttta aggtggagaa acaaatttat cagaatttcg aaaacgcgct
     5941 catcaagaag ctgaatcatt tggtcctgaa agataaagca gacgacgaaa tcggctcata
     6001 taaaaatgct ctgcagctca ctaataactt taccgacttg aaaagtatag gaaagcagac
     6061 aggctttctc ttttacgtgc ctgcttggaa cacttcaaaa atagatccag aaactggctt
     6121 tgttgacctc ctgaaaccta gatatgagaa catcgcccaa tcccaggctt tttttggcaa
     6181 gttcgacaaa atctgttaca acgccgacaa agactatttt gaattccata tagattatgc
     6241 taagtttaca gataaggcaa aaaattctag acagatatgg accatctgtt cacatgggga
     6301 caagagatac gtctatgata agactgctaa tcaaaacaaa ggagcagcta aaggtatcaa
     6361 cgtcaatgat gagctcaaaa gtttgttcgc tcgccatcac ataaacgaga agcaacctaa
     6421 tttggttatg gatatatgcc aaaacaacga taaggaattt cacaaatcgc tcatgtacct
     6481 gcttaagacg ttgctggcct tgcgctactc taacgcatcc tcagacgaag actttattct
     6541 cagtcctgtc gctaatgacg agggcgtttt tttcaactct gcgttggctg acgacacaca
     6601 accgcaaaac gcagatgcca atggcgccta ccacatcgct ctgaagggtc tttggctctt
     6661 gaatgaactc aagaacagtg atgatcttaa caaggtgaaa cttgcaatcg ataatcagac
     6721 gtggttgaat tttgcgcaga ataggaagcg tcctgctgcc accaaaaagg ccggacaggc
     6781 taagaaaaag aagtgagacg actagtggcg gccgccgacg tccgatcgtt caaacatttg
     6841 gcaataaagt ttcttaagat tgaatcctgt tgccggtctt gcgatgatta tcatataatt
     6901 tctgttgaat tacgttaagc atgtaataat taacatgtaa tgcatgacgt tatttatgag
     6961 atgggttttt atgattagag tcccgcaatt atacatttaa tacgcgatag aaaacaaaat
     7021 atagcgcgca aactaggata aattatcgcg cgcggtgtca tctatgttac tagatcggga
     7081 attgatcccc cctcgacagc ttccggaaag ggcgaattcg caactttgta tacaaaagtt
     7141 gaacgagaaa cgtaaaatga tataaatatc aatatattaa attagatttt gcataaaaaa
     7201 cagactacat aatactgtaa aacacaacat atccagtcac tatgccatcc agctgatatc
     7261 ccctatagtg agtcgtatta catggtcata gctgtttcct ggcagctctg gcccgtgtct
     7321 caaaatctct gatgttacat tgcacaagat aaaaatatat catcatgcct cctc
//
