LOCUS       Exported                2876 bp ds-DNA     circular SYN 26-FEB-2018
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pTAKN2_J23119-sgRNA
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 2876)
  AUTHORS   Nishida
  TITLE     Direct Submission
  JOURNAL   Exported Mar 2, 2018 from SnapGene 4.1.6
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..2876
                     /organism="recombinant plasmid"
                     /mol_type="other DNA"
     primer_bind     complement(21..41)
                     /label=kn696_pET-SQ2_R
     CDS             complement(230..435)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="AAAPYSESYYTSSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRN
                     SEEARTDRPSQQLRSLNGEWRL"
     primer_bind     352..375
                     /label=M13F
     primer_bind     complement(376..395)
                     /label=kn1009_M13FR
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     405..424
                     /label=T7 promoter
     promoter        405..423
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     435..477
                     /label=sb595_BamH1-EcoR1-Cla-P3gRNA_F
     CDS             complement(436..925)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="YAYLRADFCGLEKKTPPCQGRGFFFGKKAPTRCHFFKLITD*PYF
                     NLLFLALKLKSAGSSASIIPRTELAVKRKFKIIFC*LKKTPPCQGRGFFFGKKAPTRCH
                     FFKLITD*PYFNLLFLALKLRPSRVSASIIPRTELAVKRKFKIIF*CIDEDSCSIVIE"
     misc_feature    441..924
                     /label=synthetic region
     misc_feature    442..925
                     /label=insertion
     primer_bind     442..470
                     /label=sb591_Apa1-TargetRepeat_F
     primer_bind     442..470
                     /label=sb589_Sph1-TargetRepeat_F
     primer_bind     442..468
                     /label=sb602_Nar1-gRNArepeat_F
     primer_bind     442..468
                     /label=sb597_Sal1-gRNArepeat_F
     primer_bind     complement(446..470)
                     /label=kn954_OL-cI_R
     primer_bind     457..477
                     /label=sb420_Age1-upE-cnstP-1706
     primer_bind     465..550
                     /label=sb426_Cla1-upE-P3-chimrUnit_F2
     primer_bind     465..498
                     /label=sb628_BssHI-ClaI-sgRNAunit_F
     misc_feature    475..533
                     /label=promoter-Unit
     misc_feature    475..495
                     /label=up element
     promoter        499..533
                     /label=J23119
     primer_bind     499..520
                     /label=sb347_Age1-cnstPro_F
     misc_feature    499..504
                     /label=-35
     primer_bind     503..573
                     /label=sb@@@_J23119-sgRNA_F
     misc_feature    522..527
                     /label=-10
     misc_feature    534..636
                     /label=chimeric unit
     primer_bind     534..587
                     /label=kn797_chimericTracr_F
     misc_feature    534..553
                     /label=minimal CRISPR array
     misc_feature    535..540
                     /label=Bsa1
     misc_feature    547..552
                     /label=Bsa1
     primer_bind     554..576
                     /label=kn599
     misc_feature    554..568
                     /label=chimera linker
     misc_feature    569..636
                     /label=tracrRNA
     primer_bind     complement(610..629)
                     /label=kn818_tracr_R
     misc_feature    640..678
                     /label=rho-independent Ter
     primer_bind     complement(659..678)
                     /label=sb348_rep101_rhoTer_chimericUnit_R
     misc_feature    685..743
                     /label=promoter-Unit
     primer_bind     685..708
                     /label=sb628_BssHI-ClaI-sgRNAunit_F
     misc_feature    685..705
                     /label=up element
     promoter        709..743
                     /label=J23119
     primer_bind     709..730
                     /label=sb347_Age1-cnstPro_F
     misc_feature    709..714
                     /label=-35
     primer_bind     713..783
                     /label=sb@@@_J23119-sgRNA_F
     misc_feature    732..737
                     /label=-10
     misc_feature    744..846
                     /label=chimeric unit
     misc_feature    744..763
                     /label=minimal CRISPR array
     misc_feature    745..751
                     /label=Sap1
     misc_feature    756..762
                     /label=Sap1
     primer_bind     761..797
                     /label=kn797_chimericTracr_F
     primer_bind     764..786
                     /label=kn599
     misc_feature    764..778
                     /label=chimera linker
     misc_feature    779..846
                     /label=tracrRNA
     primer_bind     complement(820..839)
                     /label=kn818_tracr_R
     misc_feature    850..888
                     /label=rho-independent Ter
     primer_bind     complement(869..889)
                     /label=sb348_rep101_rhoTer_chimericUnit_R
     primer_bind     complement(889..930)
                     /label=sb596_Not1-Hind3-Xba1-P3gRNA_R
     primer_bind     complement(889..919)
                     /label=sb276_CmUnit_Xba1_F
     primer_bind     889..918
                     /label=kn992_Cm-Xba1_R
     misc_feature    895..924
                     /label=CmUnit
     primer_bind     complement(905..925)
                     /label=sb598_Pst1_gRNArepeat_R
     primer_bind     complement(906..928)
                     /label=sb592_Apa1-TargetRepeat_R
     primer_bind     complement(906..925)
                     /label=sb590_Sph1-TargetRepeat_R
     primer_bind     complement(906..924)
                     /label=sb629_BssHI-sgRNAunit_R
     primer_bind     complement(907..925)
                     /label=sb603_Aat2-gRNArepeat_R
     CDS             complement(926..929)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="S"
     primer_bind     930..949
                     /label=kn707_ColE-Bam-Spe-Sac1_R
     primer_bind     1065..1087
                     /label=kn420Gal1R-pBR322_R
     primer_bind     complement(1100..1119)
                     /label=kn746_P15AendR
     rep_origin      complement(1103..1691)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1144..1163)
                     /label=sb593_colA-ori_end_F
     primer_bind     complement(1144..1162)
                     /label=kn709_ColE1
     primer_bind     1604..1624
                     /label=sb601_ColE1ori_top_R
     primer_bind     complement(1689..1712)
                     /label=kn875_ChimerSpeNotColE_R
     primer_bind     complement(1742..1765)
                     /label=sb600_Pst1-pTAKN2-colE1-up
     primer_bind     1756..1781
                     /label=kn874_AmpAgeXbaChimr_F
     primer_bind     1766..1793
                     /label=sb599_Sal1-pTAKN2-Kan-down
     CDS             complement(1849..2664)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(1858..1881)
                     /label=kn939_Kan-end_F
     primer_bind     complement(2552..2571)
                     /label=kn887_Kan_Cla1_F
     primer_bind     2633..2648
                     /label=sb364_Km_top_R
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaatattac tacaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgagct agtgtaatac gactcactat
      421 agggcgcggc cgcagaattc gataacaatt gagcaagaat cttcatcgat gcatcagaaa
      481 attattttaa atttcctctt gacagctagc tcagtcctag gtataatgct agcagagacc
      541 cgggatggtc tcagttttag agctagaaat agcaagttaa aataaggcta gtccgttatc
      601 aacttgaaaa agtggcaccg agtcggtgct ttttttccga aaaaaaaacc ccgcccctga
      661 cagggcgggg ttttttttaa ttaacagaaa attattttaa atttcctctt gacagctagc
      721 tcagtcctag gtataatgct agcagaagag ccggcgctct tcagttttag agctagaaat
      781 agcaagttaa aataaggcta gtccgttatc aacttgaaaa agtggcaccg agtcggtgct
      841 ttttttccga aaaaaaaacc ccgcccctga cagggcgggg tttttttttc tagaccgcaa
      901 aaatcagcgc gcaaatacgc atacaagctt cctcgctcac tgactcgctg cgctcggtcg
      961 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     1021 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     1081 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     1141 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     1201 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     1261 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     1321 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     1381 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     1441 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     1501 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     1561 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     1621 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     1681 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     1741 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     1801 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgttttaatt agaaaaactc
     1861 atcgagcatc aaatgaaact gcaatttatt catatcagga ttatcaatac catatttttg
     1921 aaaaagccgt ttctgtaatg aaggagaaaa ctcaccgagg cagttccata ggatggcaag
     1981 atcctggtat cggtctgcga ttccgactcg tccaacatca atacaaccta ttaatttccc
     2041 ctcgtcaaaa ataaggttat caagtgagaa atcaccatga gtgacgactg aatccggtga
     2101 gaatggcaat agcttatgca tttctttcca gacttgttca acaggccagc cattacgctc
     2161 gtcatcaaaa tcactcgcat caaccaaacc gttattcatt cgtgattgcg cctgagcgag
     2221 acgaaatacg cgatcgctgt taaaaggaca attacaaaca ggaatcgaat gcaaccggcg
     2281 caggaacact gccagcgcat caacaatatt ttcacctgaa tcaggatatt cttctaatac
     2341 ctggaatgct gttttccctg ggatcgcagt ggtgagtaac catgcatcat caggagtacg
     2401 gataaaatgc ttgatggtcg gaagaggcat aaattccgtc agccagttta gtctgaccat
     2461 ctcatctgta acatcattgg caacgctacc tttgccatgt ttcagaaaca actctggcgc
     2521 atcgggcttc ccatacaatc gatagattgt cgcacctgat tgcccgacat tatcgcgagc
     2581 ccatttatac ccatataaat cagcatccat gttggaattt aatcgcggcc ttgagcaaga
     2641 cgtttcccgt tgaatatggc tcataacacc ccttgtatta ctgtttatgt aagcagacat
     2701 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     2761 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac
     2821 cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtc
//
