LOCUS Construct_5 10779 bp DNA circular UNA 20-APR-2016 DEFINITION U6p_20nt1_mod4_sgRNA. ACCESSION urn.local...1461188663793.13 VERSION urn.local...1461188663793.13 KEYWORDS . SOURCE Human ORGANISM Human . FEATURES Location/Qualifiers promoter 239..815 /created_by="User" /modified_by="User" /label="CMV_imm_early promoter" misc_feature 318..605 /created_by="User" /label="CAG enhancer" LTR 835..1015 /created_by="User" /modified_by="User" /label="HIV-1_5_LTR" misc_feature 1126..1170 /created_by="User" /modified_by="User" /label="HIV-1_psi_pack" misc_feature 1680..1913 /created_by="User" /label="RRE" misc_feature 2444..2459 /created_by="User" /label="cPPT" primer_bind 2594..2616 /created_by="primer3" /%GC="69.6" /Tm="64.72" /Hairpin="4.0" /Primer-Dimer="4.0" /Monovalent_Salt_Concentration="50.0" /Divalent_Salt_Concentration="0.0" /DNTP_Concentration="0.0" /Sequence="GGGGTTCCGCGCACATTTCCCCG" /Mismatches=0 /annotation_group="PrimerPlugin Group3c68cbfa-121b-4872-a7ab-08a32877e25e_urn:local:.:1365 712513487.384" /label="JP313" CDS complement(<2594..2598) /gene="AMP" /SECDrawAs="Gene" /SECStyleId=1 /label="AMP CDS" promoter 2630..>2940 /created_by="User" /modified_by="User" /label="U6 promoter" misc_feature 2727..2734 /created_by="User" /label="Enhancer" misc_feature 2883..2901 /created_by="User" /label="proximal element" misc_feature 2919..2924 /created_by="User" /label="TATA box" gRNA 2949..2967 /created_by="User" /modified_by="User" /label="20nt1" modified_base 2968..2970 /created_by="User" /modified_by="User" /label="GTT->CGG" misc_feature <2969..3046 /created_by="User" /modified_by="User" /label="sgRNA scaffold" terminator 3047..3052 /created_by="User" /label="Termination signal" CDS 3097..3612 /gene="PGKp" /SECDrawAs="Gene" /SECStyleId=1 /label="PGKp CDS" misc_feature complement(3172..3196) /Hairpin="6.0" /Primer-Dimer="2.0" /Monovalent_Salt_Concentration="50.0" /Divalent_Salt_Concentration="0.0" /DNTP_Concentration="0.0" /Mismatches=0 /created_by="primer3" /annotation_group="141566396380455900022012579-6fec-46f9-9 c44-c1191b5a2789" /%GC="68.0" /Tm="71.6" /Hairpin_Tm="64.2" /Self_Dimer_Tm="21.7" /Sequence="GGCCAGAGGCCACTTGTGTAGCGCC" /label="JP467" /note="Geneious type: primer_bind_reverse" misc_signal order(3623..3631,3623..3631) /created_by="User" /label="KOZAK" CDS 3632..4345 /gene="EBFP2" /SECDrawAs="Gene" /SECStyleId=1 /modified_by="User" /label="EBFP2 CDS" sig_peptide 4346..4411 /created_by="User" /modified_by="User" /label="p2a" CDS 4412..5443 /created_by="User" /label="hygroR" misc_feature 5473..6060 /created_by="User" /label="WPRE" misc_feature 6247..6268 /created_by="User" /modified_by="User" /label="U3PPt" misc_feature 6247..6262 /created_by="User" /label="cPPT" LTR 6584..6764 /created_by="User" /label="truncHIV-1_3_LTR" terminator 6793..7020 /created_by="User" /label="bGH_PA_terminator" rep_origin 7083..7389 /created_by="User" /label="f1_origin" misc_feature complement(7509..7724) /created_by="User" /label="SV40 enhancer" rep_origin 7688..7765 /created_by="User" /label="SV40 origin" promoter 7883..7950 /created_by="User" /label="EM7 promoter" gene 7951..8322 /created_by="User" /modified_by="User" /label="bleo" terminator 8458..8577 /created_by="User" /label="SV40_PA_terminator" promoter complement(8674..8703) /created_by="User" /modified_by="User" /label="lac_promoter" CDS complement(9786..10646) /created_by="User" /label="AmpR" promoter complement(10688..10715) /created_by="User" /label="AmpR promoter" ORIGIN 1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg 61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt 121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc 181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac 241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat 301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg 361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt 421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag 481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc 541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag 601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt 661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc 721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg 781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct 841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt 901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac 961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc 1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc 1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa 1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg 1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata 1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc 1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga 1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc 1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca 1501 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg 1561 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga 1621 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata 1681 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg 1741 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg 1801 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag 1861 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt 1921 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt 1981 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt 2041 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag 2101 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata 2161 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta 2221 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta 2281 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa 2341 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt 2401 gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat 2461 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa 2521 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag 2581 agatccagtt tggggggttc cgcgcacatt tccccgaaaa gtgccacctt gtacaaaaaa 2641 gcaggcttta aaggaaccaa ttcagtcgac tggatccggt accaaggtcg ggcaggaaga 2701 gggcctattt cccatgattc cttcatattt gcatatacga tacaaggctg ttagagagat 2761 aattagaatt aatttgactg taaacacaaa gatattagta caaaatacgt gacgtagaaa 2821 gtaataattt cttgggtagt ttgcagtttt aaaattatgt tttaaaatgg actatcatat 2881 gcttaccgta acttgaaagt atttcgattt cttggcttta tatatcttgt ggaaaggacg 2941 aaacaccgta agtcggagta ctgtcctcgg ttttagagct agaaatagca agttaaaata 3001 aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc ggtgcttttt ttctagaccc 3061 agctttcttg tacaaagttg gcattagacg tcgaggaatt ctaccgggta ggggaggcgc 3121 ttttcccaag gcagtctgga gcatgcgctt tagcagcccc gctgggcact tggcgctaca 3181 caagtggcct ctggcctcgc acacattcca catccaccgg taggcgccaa ccggctccgt 3241 tctttggtgg ccccttcgcg ccaccttcta ctcctcccct agtcaggaag ttcccccccg 3301 ccccgcagct cgcgtcgtgc aggacgtgac aaatggaagt agcacgtctc actagtctcg 3361 tgcagatgga cagcaccgct gagcaatgga agcgggtagg cctttggggc agcggccaat 3421 agcagctttg ctccttcgct ttctgggctc agaggctggg aaggggtggg tccgggggcg 3481 ggctcagggg cgggctcagg ggcggggcgg gcgcccgaag gtcctccgga ggcccggcat 3541 tctgcacgct tcaaaagcgc acgtctgccg cgctgttctc ctcttcctca tctccgggcc 3601 tttcgacctg catcccaagc ttgccaccat ggtgagcaag ggcgaggagc tgttcaccgg 3661 ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgag 3721 gggcgagggc gagggcgatg ccaccaacgg caagctgacc ctgaagttca tctgcaccac 3781 cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgagccacg gcgtgcagtg 3841 cttcgcccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga 3901 aggctacgtc caggagcgca ccatcttctt caaggacgac ggcacctaca agacccgcgc 3961 cgaggtgaag ttcgagggcg acaccctagt gaaccgcatc gagctgaagg gcgtcgactt 4021 caaggaggac ggcaacatcc tggggcacaa gctggagtac aacttcaaca gccacaacat 4081 ctatatcatg gccgtcaagc agaagaacgg catcaaggtg aacttcaaga tccgccacaa 4141 cgtggaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga 4201 cggccccgtg ctgctgcccg acagccacta cctgagcacc cagtccgtgc tgagcaaaga 4261 ccccaacgag aagcgcgatc acatggtcct gctggagttc cgcaccgccg ccgggatcac 4321 tctcggcatg gacgagctgt acaagggaag cggagctact aacttcagcc tgctgaagca 4381 ggctggagac gtggaggaga accctggacc taaaaagcct gaactcaccg cgacgtctgt 4441 cgagaagttt ctgatcgaaa agttcgacag cgtctccgac ctgatgcagc tctcggaggg 4501 cgaagaatct cgtgctttca gcttcgatgt aggagggcgt ggatatgtcc tgcgggtaaa 4561 tagctgcgcc gatggtttct acaaagatcg ttatgtttat cggcactttg catcggccgc 4621 gctcccgatt ccggaagtgc ttgacattgg ggaattcagc gagagcctga cctattgcat 4681 ctcccgccgt gcacagggtg tcacgttgca agacctgcct gaaaccgaac tgcccgctgt 4741 tctgcagccg gtcgcggagg ccatggatgc gatcgctgcg gccgatctta gccagacgag 4801 cgggttcggc ccattcggac cgcaaggaat cggtcaatac actacatggc gtgatttcat 4861 atgcgcgatt gctgatcccc atgtgtatca ctggcaaact gtgatggacg acaccgtcag 4921 tgcgtccgtc gcgcaggctc tcgatgagct gatgctttgg gccgaggact gccccgaagt 4981 ccggcacctc gtgcacgcgg atttcggctc caacaatgtc ctgacggaca atggccgcat 5041 aacagcggtc attgactgga gcgaggcgat gttcggggat tcccaatacg aggtcgccaa 5101 catcttcttc tggaggccgt ggttggcttg tatggagcag cagacgcgct acttcgagcg 5161 gaggcatccg gagcttgcag gatcgccgcg gctccgggcg tatatgctcc gcattggtct 5221 tgaccaactc tatcagagct tggttgacgg caatttcgat gatgcagctt gggcgcaggg 5281 tcgatgcgac gcaatcgtcc gatccggagc cgggactgtc gggcgtacac aaatcgcccg 5341 cagaagcgcg gccgtctgga ccgatggctg tgtagaagta ctcgccgata gtggaaaccg 5401 acgccccagc actcgtccgg atcgggagat gggggaggct aactaagaat tcgatatcaa 5461 gcttatcgat aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa 5521 ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat 5581 tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta 5641 tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc 5701 aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt 5761 ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg 5821 ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc 5881 ttggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc 5941 ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct 6001 tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgca 6061 tcgataccgt cgacctcgag acctagaaaa acatggagca atcacaagta gcaatacagc 6121 agctaccaat gctgattgtg cctggctaga agcacaagag gaggaggagg tgggttttcc 6181 agtcacacct caggtacctt taagaccaat gacttacaag gcagctgtag atcttagcca 6241 ctttttaaaa gaaaaggggg gactggaagg gctaattcac tcccaacgaa gacaagatat 6301 ccttgatctg tggatctacc acacacaagg ctacttccct gattggcaga actacacacc 6361 agggccaggg atcagatatc cactgacctt tggatggtgc tacaagctag taccagttga 6421 gcaagagaag gtagaagaag ccaatgaagg agagaacacc cgcttgttac accctgtgag 6481 cctgcatggg atggatgacc cggagagaga agtattagag tggaggtttg acagccgcct 6541 agcatttcat cacatggccc gagagctgca tccggactgt actgggtctc tctggttaga 6601 ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata 6661 aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta 6721 gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagggccc gtttaaaccc 6781 gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg 6841 tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa 6901 ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca 6961 gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg 7021 cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg 7081 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg 7141 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc 7201 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc 7261 tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga 7321 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa 7381 ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg attttgccga 7441 tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct 7501 gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat 7561 gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc 7621 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac 7681 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact 7741 aatttttttt atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta 7801 gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc 7861 cattttcgga tctgatcagc acgtgttgac aattaatcat cggcatagta tatcggcata 7921 gtataatacg acaaggtgag gaactaaacc atggccaagt tgaccagtgc cgttccggtg 7981 ctcaccgcgc gcgacgtcgc cggagcggtc gagttctgga ccgaccggct cgggttctcc 8041 cgggacttcg tggaggacga cttcgccggt gtggtccggg acgacgtgac cctgttcatc 8101 agcgcggtcc aggaccaggt ggtgccggac aacaccctgg cctgggtgtg ggtgcgcggc 8161 ctggacgagc tgtacgccga gtggtcggag gtcgtgtcca cgaacttccg ggacgcctcc 8221 gggccggcca tgaccgagat cggcgagcag ccgtgggggc gggagttcgc cctgcgcgac 8281 ccggccggca actgcgtgca cttcgtggcc gaggagcagg actgacacgt gctacgagat 8341 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc 8401 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg 8461 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa 8521 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat 8581 gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct 8641 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt 8701 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc 8761 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg 8821 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg 8881 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca 8941 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac 9001 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac 9061 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg 9121 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac 9181 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat 9241 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag 9301 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac 9361 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt 9421 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt 9481 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc 9541 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga 9601 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac 9661 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc 9721 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct 9781 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca 9841 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct 9901 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca 9961 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc 10021 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg 10081 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct 10141 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa 10201 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta 10261 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc 10321 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg 10381 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa 10441 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg 10501 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc 10561 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg 10621 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat 10681 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata 10741 ggggttccgc gcacatttcc ccgaaaagtg ccacctgac //