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x$*%  1v  qGACNNNNNNGTC,CCAGGCCTGG,GATCCGGATC,CCWGGTACCWGG,CCANNNNNTGG,CCAGGNNNTGG,CCANNNCCTGG,CCAGGNCCTGG,GCACNNNAACGTTNNNGTGC,CACNNNGTG,CCNNNNNNNGG,AACNAACRYGTGCGTGC,GCACGCACRYGTTNGTT,GCACNAACRYGTTNGTGC,AACGCACRYGTGCGTT,GACNNNNNGTC,GAANNNNNNNTTGG,CCAANNNNNNNTTC,CACNNNNGTG,AACACNNNNGTGC,GCACNNNNGTGTT,GCANNNNNNTGC,GAACNNNNNNTCC,GGANNNNNNGTTC,CAGNNNCTG,CCAGGNNCTG,CAGNNCCTGG,CCWGGGCCC,GGGCCCWGG,CCWGGGCCCWGG,AACNNNNNNGTGCACNNNNNNGTT,GACNNNGTC,GAANNNNTTC,CCWGGTACC,GGTACCWGG,CCWGGWCCWGG,ACNNNNGTAYC,GRTACNNNNGT,CCWGGYRCCWGG,GAAGNNNNNNTAC,GTANNNNNNCTTC,AACGGCNNNGTGC,GCACNNNGCCGTT,GCACGGCNNNGTT,AACNNNGCCGTGC,CGANNNNNNTGC,GCANNNNNNTCG,CGATCNNNNTGC,GCANNNNGATCG,TGANNNNNNTCA,TGATCNNNNTCA,TGANNNNGATCA,TGATCNNGATCA,AACCTGCNNGTGC,GCACNNGCAGGTT,GCACCTGCNNGTT,AACNNGCAGGTGC,GCCNNNNNGGC,AACNNNNNNGTGCMC,GKGCACNNNNNNGTT,CCWGGNCCWGG,AACTGGGNNGTGC,GCACNNCCCAGTT,GCACTGGGNNGTT,AACNNCCCAGTGC,GACNNNNGTC,GAGNNNNNCTC,CCWGGTCTC,GAGACCWGG,GATNNNNATC,GATCNNNATC,GATNNNGATC,GATCNNGATC,CCWGGCGCCWGG,ACNNNNNCTCC,GGAGNNNNNGT,AACNNNNNNGTGCAG,CTGCACNNNNNNGTT,GATCATGATC,CCWGGNNCCWGG,AACTGGNNNGTGC,GCACNNNCCAGTT,GCACTGGNNNGTT,AACNNNCCAGTGC,AACNAACCGGTGCGTGC,GCACGCACCGGTTNGTT,GCACNAACCGGTTNGTGC,AACGCACCGGTGCGTT,GCANNNNNTGC,GCACNNNNNNGTTCGAACNNNNNNGTGC,CCTNNNNNAGG,GCNNNNNNNGC,CCANNNNNNTGG,CCAGGNNCCTGG,CCTGGCCAGG,CAANNNNNGTGG,CCACNNNNNTTG,TTTAAACNNNNNNGTGC,GCACNNNNNNGTTTAAA,GCACNNNNNNGTTTAAACNNNNNNGTGC,CCAGGNCCY,RGGNCCTGG,CCWGGGNCCTGG,CCAGGNCCCWGG,AAGNNNNNCTT,GAYNNNNNVTC,GABNNNNNRTC,GAYNNNNNRTC,GATCNNNNVTC,GABNNNNGATC,GAYNNNNGATC,GATCNNNNRTC,GATCNNNGATC,GTYAACNNNNNNGTGC,GCACNNNNNNGTTRAC,GCACNNNNNNGTTAACNNNNNNGTGC,GATCNGATC,TAACTATAACGGTCCTAAGGTAGCGA,TCGCTACCTTAGGACCGTTATAGTTA,CTCTCTTAAGGTAGC,GCTACCTTAAGAGAG,TAGGGATAACAGGGTAAT,ATTACCCTGTTATCCCTA,GTTAACNNNNNNGTGC,GCACNNNNNNGTTAAC,AACNAACGCGTGCGTGC,GCACGCACGCGTTNGTT,GCACNAACGCGTTNGTGC,AACGCACGCGTGCGTT,TCCAACNNNNNNGTGC,GCACNNNNNNGTTGGA,CAYNNNNRTG,AACAYNNNNGTGC,GCACNNNNRTGTT,GTTTAAACNNNNNNGTGC,GCACNNNNNNGTTTAAAC,CCWGGCGCC,GGCGCCWGG,CCWGGNNCC,GGNNCCWGG,GATCGCGATC,AACNAACATGTGCGTGC,GCACGCACATGTTNGTT,GCACNAACATGTTNGTGC,AACGCACATGTGCGTT,TGGCAAACAGCTATTATGGGTATTATGGGT,ACCCATAATACCCATAATAGCTGTTTGCCA,ATCTATGTCGGGTGCGGAGAAAGAGGTAATGAAATGG,CCATTTCATTACCTCTTTCTCCGCACCCGACATAGAT,AACNNGCACGTGCNNGTT,GAACNNNNNCTC,GAGNNNNNGTTC,CCAGGWCCY,RGGWCCTGG,CCWGGGWCCTGG,CCAGGWCCCWGG,CCAGGWCCTGG,TTATAACNNNNNNGTGC,GCACNNNNNNGTTATAA,GCACNNNNNNGTTATAACNNNNNNGTGC,GAACNNNNNNTAC,GTANNNNNNGTTC,GGCCNNNNNGGCC,AACNNACTAGTGC,GCACTAGTNNGTT,AACTAGTNNGTGC,GCACNNACTAGTT,AACNAACTAGTGCGTGC,GCACGCACTAGTTNGTT,GCACNAACTAGTTNGTGC,AACGCACTAGTGCGTT,AACNNNAACGTGCNNGTGC,GCACNNGCACGTTNNNGTT,GCACCCANNNGTT,AACNNNTGGGTGC,TTAACNNNNNNGTGC,GCACNNNNNNGTTAA,CACNNNNNNTCC,GGANNNNNNGTG,CACNNNNGATCC,GGATCNNNNGTG,GATCTAGATC,CCANNNNNNNNNTGG,CCAGGCCT,AGGCCTGG,CCTCGAGG,GATCCGGA,TCCGGATC,GGCGCGCC,GCGATCGC,CCWGGWCC,GGWCCWGG,GATCGATC,GATCATGA,TCATGATC,GCGGCCGC,CCTGGCCR,YGGCCAGG,CCWGGNCC,GGNCCWGG,GGCCGGCC,RTGCGCAY,GATCNNGA,TCNNGATC,CGCGCGCG,CCTGGCCA,TGGCCAGG,CGCCGGCG,GTTTAAAC,GATCGCGA,TCGCGATC,TTAATTAA,VCTCGAGB,CCTGCAGG,CRCCGGYG,CGTCGACG,ATTTAAAT,GCCCGGGC,GATCTAGA,TCTAGATC,CACCTGC,GCAGGTG,CCTNAGG,GATCGAT,ATCGATC,CCTCAGC,GCTGAGG,GCTNAGC,CCTNAGC,GCTNAGG,GCTCTTC,GAAGAGC,GGTNACC,CGGWCCG,ACCWGGT,RGGNCCY,CCTGGAG,CTCCAGG,GGTGATC,GATCACC,GATCNGA,TCNGATC,GGGWCCC,GAAGATC,GATCTTC,CCCWGGG,TCCNGGA,TCCWGGA,RGGWCCY,TTATAA,AGGCCT,GACGTC,GTMKAC,TCCGGA,TGCGCA,ACCTGC,GCAGGT,GGTACC,AGTACT,GGYRCC,CCGCTC,GAGCGG,AACGTT,YGGCCR,RAATTY,CTGAAG,CTTCAG,CACGTG,GRCGYC,AGCGCT,CTTAAG,ACRYGT,ACCGGT,ACTAGT,CACGTC,GACGTG,GWGCWC,GTGCAC,CYCGRG,GGGCCC,ATTAAT,CCTAGG,TTCGAA,GCTAGC,GKGCMC,TGGCCA,GGATCC,GRGCYC,ATCGAT,CACGAG,CTCGTG,GGCGCC,GAAGAC,GTCTTC,GCATGC,GTATCC,GGATAC,TGATCA,ACTGGG,CCCAGT,CTRYAG,RGCGCY,AGATCT,GGNNCC,CTGGAG,CTCCAG,CTTGAG,CTCAAG,CGATCG,GGTCTC,GAGACC,YACGTR,CCNNGG,GAATGC,GCATTC,WCCGGW,GCAATG,CATTGC,RCCGGY,GCGCGC,GAGGAG,CTCCTC,CGGCCG,CCCAGC,GCTGGG,GTGCAG,CTGCAC,CGRYCG,CGTACG,CGTCTC,GAGACG,CCATGG,TCGCGA,GDGCHC,TGTACA,TCATGA,ACATGT,GTATAC,CCWWGG,CTCTTC,GAAGAG,GCNNGC,CCRYGG,GTTAAC,RCATGY,TACGTA,RGATCY,GCGATG,CATCGC,GCAGTG,CACTGC,CCCGGG,CCGCGG,TTTAAA,GGCGGA,TCCGCC,GAGCTC,GATATC,CTGRAG,CTYCAG,CAGCAG,CTGCTG,GAATTC,ATGCAT,CATATG,GTYRAC,AAGCTT,GTNNAC,TCNNGA,CAATTG,ACGCGT,TCCRAC,GTYGGA,GCCGGC,CMGCKG,GCCGAG,CTCGGC,CTCGAG,CTGCAG,CAGCTG,GTCGAC,CTYRAG,AATATT,GATCGA,TCGATC,GACCGA,CACCCA,TCGGTC,TGGGTG,WGTACW,TARCCA,TGGYTA,TCTAGA,GGATC,GATCC,CCWGG,GTCTC,GAGAC,GCWGC,GGNCC,CCSGG,GGTGA,TCACC,GGWCC,GCAGC,GCTGC,CCATC,GATGG,ACGGC,GCCGT,GCNGC,CCNGG,GCATC,GATGC,ACTGG,CCAGT,GGATG,CATCC,CTCAG,CTGAG,GGGAC,GTCCC,ACNGT,CTNAG,CAGTG,CACTG,GACGC,GCGTC,CCCGC,GCGGG,GANTC,CGWCG,TCNGA,CCTTC,GAAGG,GTNAC,GAAGA,TCTTC,GAGTC,GACTC,GTSAC,GAWTC,GCSGC,CASTG,ATGAA,TTCAT,ACGGA,TCCGT,CGCG,CCGC,GCGG,GTAC,AGCT,GCGC,CTAG,GATC,GGCC,CCGG,CATG,RGCY,YATR,ACGT,TGCA,AATT,CCTC,GAGG,TTAA,ASST,TCGA                                                                                                                                                                                                                                                                                                                                                       3                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                	                                                                                                                                                                                                                                                                      ,                	                      
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the &lt;i&gt;araC&lt;/i&gt; regulatory gene is transcribed in the opposite direction (&lt;a href=&quot;http://www.ncbi.nlm.nih.gov/pubmed/7608087&quot; title=&quot;http://www.ncbi.nlm.nih.gov/pubmed/7608087&quot;&gt;Guzman et al., 1995&lt;/a&gt;)&lt;/body&gt;&lt;/html&gt;"/></Q></Feature><Feature recentID="7" name="pSC101 ori" type="rep_origin" swappedSegmentNumbering="1" allowSegmentOverlaps="0" consecutiveTranslationNumbering="1"><Segment range="2821-3043" color="#ffff00" type="standard"/><Q name="note"><V text="&lt;html&gt;&lt;body&gt;low-copy replication origin that requires the Rep101 protein&lt;/body&gt;&lt;/html&gt;"/></Q></Feature><Feature recentID="8" name="Rep101(Ts)" directionality="2" translationMW="37384.08" type="CDS" swappedSegmentNumbering="1" allowSegmentOverlaps="0" readingFrame="-1" consecutiveTranslationNumbering="1" hitsStopCodon="1"><Segment range="1823-2773" color="#993366" type="standard" translated="1"/><Q name="codon_start"><V int="1"/></Q><Q name="gene"><V text="&lt;html&gt;&lt;body&gt;&lt;i&gt;rep101&lt;/i&gt; encoding an A56V mutation&lt;/body&gt;&lt;/html&gt;"/></Q><Q name="product"><V text="&lt;html&gt;&lt;body&gt;temperature-sensitive version of the RepA protein needed for replication with the pSC101 origin (&lt;a href=&quot;http://www.ncbi.nlm.nih.gov/pubmed/6327996&quot; title=&quot;http://www.ncbi.nlm.nih.gov/pubmed/6327996&quot;&gt;Armstrong et al., 1984&lt;/a&gt;)&lt;/body&gt;&lt;/html&gt;"/></Q><Q name="transl_table"><V int="1"/></Q><Q name="translation"><V text="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKERTVSFTYNQYVQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFSSEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANIEISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQVELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKFLSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI*"/></Q></Feature><Feature recentID="9" name="Shine-Dalgarno sequence" type="RBS" allowSegmentOverlaps="0" consecutiveTranslationNumbering="1"><Segment range="7987-7995" color="#a6acb3" type="standard"/><Q name="note"><V text="&lt;html&gt;&lt;body&gt;full consensus sequence for ribosome-binding sites upstream of start codons in &lt;i&gt;E. coli&lt;/i&gt;; complementary to a region in the 3' end of the 16S rRNA (&lt;a href=&quot;https://www.ncbi.nlm.nih.gov/pubmed/7528374&quot; title=&quot;https://www.ncbi.nlm.nih.gov/pubmed/7528374&quot;&gt;Chen et al., 1994&lt;/a&gt;)&lt;/body&gt;&lt;/html&gt;"/></Q></Feature><Feature recentID="10" name="Gam" directionality="1" translationMW="16346.73" type="CDS" allowSegmentOverlaps="0" consecutiveTranslationNumbering="1" hitsStopCodon="1"><Segment range="8004-8420" color="#ff0000" type="standard" translated="1"/><Q name="codon_start"><V int="1"/></Q><Q name="gene"><V text="&lt;html&gt;&lt;body&gt;&lt;i&gt;gam&lt;/i&gt;&lt;/body&gt;&lt;/html&gt;"/></Q><Q name="note"><V text="&lt;html&gt;&lt;body&gt;protects the double-stranded DNA substrate for recombination&lt;/body&gt;&lt;/html&gt;"/></Q><Q name="product"><V text="&lt;html&gt;&lt;body&gt;inhibitor of the host RecBCD nuclease in the&amp;nbsp;Î» Red system&lt;/body&gt;&lt;/html&gt;"/></Q><Q name="transl_table"><V int="1"/></Q><Q name="translation"><V text="MDINTETEIKQKHSLTPFPVFLISPAFRGRYFHSYFRSSAMNAYYIQDRLEAQSWARHYQQLAREEKEAELADDMEKGLPQHLFESLCIDHLQRHGASKKSITRAFDDDVEFQERMAEHIRYMVETIAHHQVDIDSEV*"/></Q></Feature><Feature recentID="11" name="Beta" directionality="1" translationMW="29689.57" type="CDS" allowSegmentOverlaps="0" consecutiveTranslationNumbering="1" hitsStopCodon="1"><Segment range="8426-9211" color="#ff0000" type="standard" translated="1"/><Q name="codon_start"><V int="1"/></Q><Q name="gene"><V text="&lt;html&gt;&lt;body&gt;&lt;i&gt;bet&lt;/i&gt;&lt;/body&gt;&lt;/html&gt;"/></Q><Q name="note"><V text="&lt;html&gt;&lt;body&gt;mediates pairing of a single-stranded DNA with a complementary target&lt;/body&gt;&lt;/html&gt;"/></Q><Q name="product"><V text="&lt;html&gt;&lt;body&gt;single-stranded DNA binding recombinase in the&amp;nbsp;Î» Red system&lt;/body&gt;&lt;/html&gt;"/></Q><Q name="transl_table"><V int="1"/></Q><Q name="translation"><V text="MSTALATLAGKLAERVGMDSVDPQELITTLRQTAFKGDASDAQFIALLIVANQYGLNPWTKEIYAFPDKQNGIVPVVGVDGWSRIINENQQFDGMDFEQDNESCTCRIYRKDRNHPICVTEWMDECRREPFKTREGREITGPWQSHPKRMLRHKAMIQCARLAFGFAGIYDKDEAERIVENTAYTAERQPERDITPVNDETMQEINTLLIALDKTWDDDLLPLCSQIFRRDIRASSELTQAEAVKALGFLKQKAAEQKVAA*"/></Q></Feature><Feature recentID="12" name="Exo" directionality="1" translationMW="25909.26" type="CDS" allowSegmentOverlaps="0" consecutiveTranslationNumbering="1" hitsStopCodon="1"><Segment range="9208-9888" color="#ff0000" type="standard" translated="1"/><Q name="codon_start"><V int="1"/></Q><Q name="gene"><V text="&lt;html&gt;&lt;body&gt;&lt;i&gt;exo&lt;/i&gt;&lt;/body&gt;&lt;/html&gt;"/></Q><Q name="note"><V text="&lt;html&gt;&lt;body&gt;creates 3' single-stranded DNA tails&lt;/body&gt;&lt;/html&gt;"/></Q><Q name="product"><V text="&lt;html&gt;&lt;body&gt;5' to 3' double-stranded DNA exonuclease in the&amp;nbsp;Î» Red system&lt;/body&gt;&lt;/html&gt;"/></Q><Q name="transl_table"><V int="1"/></Q><Q name="translation"><V text="MTPDIILQRTGIDVRAVEQGDDAWHKLRLGVITASEVHNVIAKPRSGKKWPDMKMSYFHTLLAEVCTGVAPEVNAKALAWGKQYENDARTLFEFTSGVNVTESPIIYRDESMRTACSPDGLCSDGNGLELKCPFTSRDFMKFRLGGFEAIKSAYMAQVQYSMWVTRKNAWYFANYDPRMKREGLHYVVIERDEKYMASFDEIVPEFIEKMDEALAEIGFVFGEQWR*"/></Q></Feature><Feature recentID="13" name="Î» tL3 terminator" type="terminator" swappedSegmentNumbering="1" allowSegmentOverlaps="0" readingFrame="-1" consecutiveTranslationNumbering="1"><Segment range="9889-10133" color="#ffffff" type="standard"/><Q name="note"><V text="&lt;html&gt;&lt;body&gt;transcription terminator &lt;i&gt;t&lt;/i&gt;&lt;sub&gt;L3&lt;/sub&gt; from phage&amp;nbsp;Î»&lt;/body&gt;&lt;/html&gt;"/></Q></Feature></Features>
  ç<?xml version="1.0"?><Primers nextValidID="4"><HybridizationParams minContinuousMatchLen="10" allowMismatch="1" minMeltingTemperature="40"/><Primer recentID="0" name="Kan-R" sequence="ATCGCGAGCCCATTTATACC" description="Kanamycin resistance gene, reverse primer"><BindingSite location="4417-4436" boundStrand="0" annealedBases="ATCGCGAGCCCATTTATACC" meltingTemperature="57"><Component hybridizedRange="4417-4436" bases="ATCGCGAGCCCATTTATACC"/></BindingSite><BindingSite simplified="1" location="4417-4436" boundStrand="0" annealedBases="ATCGCGAGCCCATTTATACC" meltingTemperature="57"><Component hybridizedRange="4417-4436" bases="ATCGCGAGCCCATTTATACC"/></BindingSite></Primer><Primer recentID="1" name="LacI-R" sequence="GGCATACTCTGCGACATCGT" description="LacI, reverse primer"><BindingSite location="1573-1592" boundStrand="0" annealedBases="GGCATACTCTGCGACATCGT" meltingTemperature="59"><Component hybridizedRange="1573-1592" bases="GGCATACTCTGCGACATCGT"/></BindingSite><BindingSite simplified="1" location="1573-1592" boundStrand="0" annealedBases="GGCATACTCTGCGACATCGT" meltingTemperature="59"><Component hybridizedRange="1573-1592" bases="GGCATACTCTGCGACATCGT"/></BindingSite></Primer><Primer recentID="2" name="pBAD Forward" sequence="ATGCCATAGCATTTTTATCC" description="For vectors with E. coli araBAD promoter, forward primer"><BindingSite location="7882-7901" boundStrand="0" annealedBases="ATGCCATAGCATTTTTATCC" meltingTemperature="51"><Component hybridizedRange="7882-7901" bases="ATGCCATAGCATTTTTATCC"/></BindingSite><BindingSite simplified="1" location="7882-7901" boundStrand="0" annealedBases="ATGCCATAGCATTTTTATCC" meltingTemperature="51"><Component hybridizedRange="7882-7901" bases="ATGCCATAGCATTTTTATCC"/></BindingSite></Primer><Primer recentID="3" name="pENTR-R" sequence="ATGGCTCATAACACCCCTTG" description="pENTR vectors, reverse primer"><BindingSite location="4501-4520" boundStrand="0" annealedBases="ATGGCTCATAACACCCCTTG" meltingTemperature="57"><Component hybridizedRange="4501-4520" bases="ATGGCTCATAACACCCCTTG"/></BindingSite><BindingSite simplified="1" location="4501-4520" boundStrand="0" annealedBases="ATGGCTCATAACACCCCTTG" meltingTemperature="57"><Component hybridizedRange="4501-4520" bases="ATGGCTCATAACACCCCTTG"/></BindingSite></Primer></Primers>
  
<Notes>
<UUID>ef07b427-d102-484c-8421-a99af0c577b3</UUID>
<Type>Synthetic</Type>
<ConfirmedExperimentally>0</ConfirmedExperimentally>
<CustomMapLabel>pCas12f1</CustomMapLabel>
<UseCustomMapLabel>1</UseCustomMapLabel>
<Description>Constitutive expression of cas12f1, arabinose-inducible expression of lambda recombinase, and IPTG-inducible expression of sgRNA for curing of pTarget12f1.</Description>
<Created UTC="19:36:21">2021.6.29</Created>
<LastModified UTC="19:36:21">2021.6.29</LastModified>
<SequenceClass>UNA</SequenceClass>
<TransformedInto>unspecified</TransformedInto>
<References>
<Reference journalName="Unpublished" journal="Unpublished" type="Journal Article" title="Genome editing by miniature CRISPR/Cas12f1 enzyme in Escherichia coli"/>
</References>
</Notes>
  Y     –         
?????                                                                                                       ÿ d      HO  d               E              ÿÿÿÿYô? P                                                                                                                                                              3<?xml version="1.0"?><EnzymeVisibilities vals=""/>
   )<?xml version="1.0"?><CustomEnzymeSets/>
