LOCUS       pFH86                   2878 bp    DNA     circular UNA 03-JUN-2019
DEFINITION  Subcloning of protospacer into SpCas9 improved sgRNAscaffold,
            overhang to pFH33 TaU3p.
ACCESSION   urn.local...1m-ajo6shz
VERSION     urn.local...1m-ajo6shz
KEYWORDS    .
SOURCE      
  ORGANISM  
            .
FEATURES             Location/Qualifiers
     misc_feature    2..7
                     /Recognition_pattern="GGTCTC(1/5)"
                     /annotation_group="GGTCTC"
                     /label="BsaI"
                     /note="Geneious type: restriction site"
     overhang        9..12
                     /created_by="hahnf"
                     /label="AGCA to TaU3p pFH33"
     misc_feature    complement(15..20)
                     /Recognition_pattern="GAAGAC(2/6)"
                     /annotation_group="GAAGAC"
                     /label="BpiI"
                     /note="Geneious type: restriction site"
     CDS             21..598
                     /created_by="hahnf"
                     /label="lacZ"
     misc_feature    599..604
                     /Recognition_pattern="GAAGAC(2/6)"
                     /annotation_group="GAAGAC"
                     /label="BpiI"
                     /note="Geneious type: restriction site"
     gRNA_Scaffold   607..692
                     /created_by="hahnf"
                     /modified_by="hahnf"
                     /label="SpCas9 improved sgRNA scaffold (Dang)"
     overhang        607..610
                     /created_by="hahnf"
                     /modified_by="hahnf"
                     /label="GTTT"
     Stop_codon      693..701
                     /created_by="hahnf"
                     /label="STOP"
     overhang        702..705
                     /created_by="hahnf"
                     /modified_by="hahnf"
                     /label="CGCT overhang to Level1 backbone"
     misc_feature    complement(707..712)
                     /Recognition_pattern="GGTCTC(1/5)"
                     /annotation_group="GGTCTC"
                     /label="BsaI"
                     /note="Geneious type: restriction site"
     rep_origin      complement(943..1557)
                     /vntifkey=21
                     /X-Transferred_From="pAK-EL-01; pAK-EL-02; pAK-EL-03;
                     pAK-EL-04; pAK-EL-05"
                     /X-Transferred_Similarity="100.00%"
                     /Transferred_From="pAK-EL-01, pAK-EL-02, pAK-EL-03,
                     pAK-EL-04, pAK-EL-05, pAK007 in pUC57-Kan-mini, pAK008 in
                     pUC57-Kan-mini, pAK009 in pUC57-Kan-mini, pAK010 in
                     pUC57-Kan-mini, pAK011 in pUC57-Kan-mini, pAK012 in
                     pUC57-Kan-mini"
                     /Transferred_Similarity="100.00%"
                     /label="rep(pMB1)"
     CDS             complement(1682..2497)
                     /vntifkey=21
                     /X-Transferred_From="pAK-EL-01; pAK-EL-02; pAK-EL-03;
                     pAK-EL-04; pAK-EL-05"
                     /X-Transferred_Similarity="100.00%"
                     /Transferred_From="pAK-EL-01, pAK-EL-02, pAK-EL-03,
                     pAK-EL-04, pAK-EL-05, pAK007 in pUC57-Kan-mini, pAK008 in
                     pUC57-Kan-mini, pAK009 in pUC57-Kan-mini, pAK010 in
                     pUC57-Kan-mini, pAK011 in pUC57-Kan-mini, pAK012 in
                     pUC57-Kan-mini"
                     /Transferred_Similarity="98.90%"
                     /label="KanR"
     primer_bind     2568..2591
                     /Mismatches=0
                     /%GC="33.3"
                     /Hairpin_Tm="None"
                     /Self_Dimer_Tm="None"
                     /Tm="56.0"
                     /annotation_group="245561209309456a9bc8b4-644a-4aa5-8b92-f
                     e3200f2d5ce"
                     /Sequence="GCAATGTAACATCAGAGATTTTGA"
                     /created_by="primer3"
                     /label="FH32"
ORIGIN      
        1 gggtctcaag cattgtcttc gcagctggca cgacaggttt cccgactgga aagcgggcag
       61 tgagcgcaac gcaattaatg tgagttagct cactcattag gcaccccagg ctttacactt
      121 tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc acacaggaaa
      181 cagctatgac catgattacg ccaagcttgc atgcctgcag gtcgactcta gaggatcccc
      241 gggtaccgag ctcgaattca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg
      301 gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg
      361 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggcgcc
      421 tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata tggtgcactc
      481 tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg
      541 ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgacga
      601 agacaagttt cagagctatg ctggaaacag catagcaagt tgaaataagg ctagtccgtt
      661 atcaacttga aaaagtggca ccgagtcggt gctttttttt tcgcttgaga ccgaggctag
      721 gtggaggctc agtgatgata agtctgcgat ggtggatgca tgtgtcatgg tcatagctgt
      781 ttcctgtgtg aaattgttat ccgctcagag ggcacaatcc tattccgcgc tatccgacaa
      841 tctccaagac attaggtgga gttcagttcg gcgtatggca tatgtcgctg gaaagaacat
      901 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
      961 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     1021 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     1081 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     1141 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     1201 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     1261 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     1321 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     1381 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     1441 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     1501 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     1561 cttttctacg gggtctgacg ctctattcaa caaagccgcc gtcccgtcaa gtcagcgtaa
     1621 atgggtaggg ggcttcaaat cgtccgctct gccagtgtta caaccaatta acaaattctg
     1681 attagaaaaa ctcatcgagc atcaaatgaa actgcaattt attcatatca ggattatcaa
     1741 taccatattt ttgaaaaagc cgtttctgta atgaaggaga aaactcaccg aggcagttcc
     1801 ataggatggc aagatcctgg tatcggtctg cgattccgac tcgtccaaca tcaatacaac
     1861 ctattaattt cccctcgtca aaaataaggt tatcaagtga gaaatcacca tgagtgacga
     1921 ctgaatccgg tgagaatggc aaaagcttat gcatttcttt ccagacttgt tcaacaggcc
     1981 agccattacg ctcgtcatca aaatcactcg catcaaccaa accgttattc attcgtgatt
     2041 gcgcctgagc gagacgaaat acgcgatcgc tgttaaaagg acaattacaa acaggaatcg
     2101 aatgcaaccg gcgcaggaac actgccagcg catcaacaat attttcacct gaatcaggat
     2161 attcttctaa tacctggaat gctgttttcc cggggatcgc agtggtgagt aaccatgcat
     2221 catcaggagt acggataaaa tgcttgatgg tcggaagagg cataaattcc gtcagccagt
     2281 ttagtctgac catctcatct gtaacatcat tggcaacgct acctttgcca tgtttcagaa
     2341 acaactctgg cgcatcgggc ttcccataca atcgatagat tgtcgcacct gattgcccga
     2401 cattatcgcg agcccattta tacccatata aatcagcatc catgttggaa tttaatcgcg
     2461 gcctcgagca agacgtttcc cgttgaatat ggctcataac accccttgta ttactgttta
     2521 tgtaagcaga cagttttatt gttcatgatg atatattttt atcttgtgca atgtaacatc
     2581 agagattttg agacacaacg tggctttccc ccgccgctct agaactagtg gatccaaata
     2641 aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta tctgttgttt gtcgcattat
     2701 acgagacgtc caggttggga tacctgaaac aaaacccatc gtacggccaa ggaagtctcc
     2761 aataactgtg atccaccaca agcgccaggg ttttcccagt cacgacgttg taaaacgacg
     2821 gccagtcatg cataatccgc acgcatctgg aataaggaag tgccattccg cctgacct
//
