LOCUS       Cas9_mCherry_HSF2_KO    9721 bp ds-DNA     circular     15-MAR-2023
DEFINITION  .
FEATURES             Location/Qualifiers
     rep_origin      60..648
                     /label="ori"
                     /ApEinfo_revcolor="#b1ff67"
                     /ApEinfo_fwdcolor="#b1ff67"
     primer_bind     549..568
                     /label="pBR322ori-F"
                     /ApEinfo_revcolor="#f58a5e"
                     /ApEinfo_fwdcolor="#f58a5e"
     primer          complement(695..714)
                     /label="oST060"
                     /note="sequence: ccctcacatgtgagcaaaag"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
     misc_feature    710..950
                     /label="U6 Promoter"
                     /ApEinfo_revcolor="#9eafd2"
                     /ApEinfo_fwdcolor="#9eafd2"
     misc_feature    1421..1496
                     /label="RNA Scafhold"
                     /ApEinfo_revcolor="#ff9ccd"
                     /ApEinfo_fwdcolor="#ff9ccd"
     misc_feature    1593..1878
                     /label="CMV Enhancer"
                     /ApEinfo_revcolor="#d59687"
                     /ApEinfo_fwdcolor="#d59687"
     promoter        1880..2157
                     /label="chicken beta-actin promoter"
                     /ApEinfo_revcolor="#d59687"
                     /ApEinfo_fwdcolor="#d59687"
     intron          2158..2386
                     /label="hybrid intron"
                     /ApEinfo_revcolor="#ffef86"
                     /ApEinfo_fwdcolor="#ffef86"
     regulatory      2398..2407
                     /label="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)"
                     /ApEinfo_revcolor="#c7b0e3"
                     /ApEinfo_fwdcolor="#c7b0e3"
     CDS             2407..2472
                     /label="3xFLAG"
                     /ApEinfo_revcolor="#d59687"
                     /ApEinfo_fwdcolor="#d59687"
     CDS             2479..2499
                     /label="SV40 NLS"
                     /ApEinfo_revcolor="#d59687"
                     /ApEinfo_fwdcolor="#d59687"
     CDS             2524..6624
                     /label="Cas9"
                     /ApEinfo_revcolor="#faac61"
                     /ApEinfo_fwdcolor="#faac61"
     CDS             6625..6672
                     /label="nucleoplasmin NLS"
                     /ApEinfo_revcolor="#ff9ccd"
                     /ApEinfo_fwdcolor="#ff9ccd"
     CDS             6676..6729
                     /label="T2A"
                     /ApEinfo_revcolor="#c7b0e3"
                     /ApEinfo_fwdcolor="#c7b0e3"
     CDS             6736..7446
                     /label="mCherry"
                     /ApEinfo_revcolor="#faac61"
                     /ApEinfo_fwdcolor="#faac61"
     primer_bind     complement(6881..6899)
                     /label="mCherry-R"
                     /ApEinfo_revcolor="#d6b295"
                     /ApEinfo_fwdcolor="#d6b295"
     primer_bind     complement(7471..7488)
                     /label="BGH-rev"
                     /ApEinfo_revcolor="#ff9ccd"
                     /ApEinfo_fwdcolor="#ff9ccd"
     polyA_signal    7477..7684
                     /label="bGH poly(A) signal"
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
     repeat_region   7693..7822
                     /label="AAV2 ITR"
                     /ApEinfo_revcolor="#d59687"
                     /ApEinfo_fwdcolor="#d59687"
     repeat_region   7693..7833
                     /label="AAV2 ITR"
                     /ApEinfo_revcolor="#ff9ccd"
                     /ApEinfo_fwdcolor="#ff9ccd"
     rep_origin      7908..8363
                     /label="f1 ori"
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
     primer_bind     complement(7995..8014)
                     /label="F1ori-R"
                     /ApEinfo_revcolor="#b1ff67"
                     /ApEinfo_fwdcolor="#b1ff67"
     primer_bind     8205..8226
                     /label="F1ori-F"
                     /ApEinfo_revcolor="#faac61"
                     /ApEinfo_fwdcolor="#faac61"
     primer_bind     complement(8380..8399)
                     /label="pRS-marker"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
     primer_bind     8499..8521
                     /label="pGEX 3'"
                     /ApEinfo_revcolor="#85dae9"
                     /ApEinfo_fwdcolor="#85dae9"
     primer_bind     complement(8559..8577)
                     /label="pBRforEco"
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
     promoter        8645..8749
                     /label="AmpR promoter"
                     /ApEinfo_revcolor="#75c6a9"
                     /ApEinfo_fwdcolor="#75c6a9"
     primer          8690..8719
                     /label="oST795"
                     /note="sequence: TATCCGCTCATGAGACAATAACCCTGATAA"
                     /ApEinfo_revcolor="#faac61"
                     /ApEinfo_fwdcolor="#faac61"
     CDS             8750..9610
                     /label="AmpR"
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
     primer_bind     complement(8968..8987)
                     /label="Amp-R"
                     /ApEinfo_revcolor="#f58a5e"
                     /ApEinfo_fwdcolor="#f58a5e"
ORIGIN
        1 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
       61 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
      121 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
      181 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
      241 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
      301 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
      361 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
      421 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
      481 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
      541 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
      601 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaAA CGCCAGCAAC
      661 GCGGCCTTTT TACGGTTCCT GGCCTTTTGC TGGCCTTTTG CTCACATGTG AGGGCCTATT
      721 TCCCATGATT CCTTCATATT TGCATATACG ATACAAGGCT GTTAGAGAGA TAATTGGAAT
      781 TAATTTGACT GTAAACACAA AGATATTAGT ACAAAATACG TGACGTAGAA AGTAATAATT
      841 TCTTGGGTAG TTTGCAGTTT TAAAATTATG TTTTAAAATG GACTATCATA TGCTTACCGT
      901 AACTTGAAAG TATTTCGATT TCTTGGCTTT ATATATCTTG TGGAAAGGAC GAAACACCGa
      961 attctactac cgaacgcggG TTTTAGAGCT AGAAATAGCA AGTTAAAATA AGGCTAGTCC
     1021 GTTATCAACT TGAAAAAGTG GCACCGAGTC GGTGCTTTTT TTCTAGACCC AGCTTTCTTG
     1081 TTGTACAAAA AAGCAGGCTT TAAAGGAACC AATTCAGTCG ACTGGATCCG GTACCAAGGT
     1141 CGGGCAGGAA GAGGGCCTAT TTCCCATGAT TCCTTCATAT TTGCATATAC GATACAAGGC
     1201 TGTTAGAGAG ATAATTAGAA TTAATTTGAC TGTAAACACA AAGATATTAG TACAAAATAC
     1261 GTGACGTAGA AAGTAATAAT TTCTTGGGTA GTTTGCAGTT TTAAAATTAT GTTTTAAAAT
     1321 GGACTATCAT ATGCTTACCG TAACTTGAAA GTATTTCGAT TTCTTGGCTT TATATATCTT
     1381 GTGGAAAGGA CGAAACACCG tccaagccag gtaggaatgg GTTTTAGAGC TAGAAATAGC
     1441 AAGTTAAAAT AAGGCTAGTC CGTTATCAAC TTGAAAAAGT GGCACCGAGT CGGTGCTTTT
     1501 TTGTTTTAGA GCTAGAAATA GCAAGTTAAA ATAAGGCTAG TCCGTTTTTA GCGCGTGCGC
     1561 CAATTCTGCA GACAAATGGC TCTAGAGGTA CCCGTTACAT AACTTACGGT AAATGGCCCG
     1621 CCTGGCTGAC CGCCCAACGA CCCCCGCCCA TTGACGTCAA TAGTAACGCC AATAGGGACT
     1681 TTCCATTGAC GTCAATGGGT GGAGTATTTA CGGTAAACTG CCCACTTGGC AGTACATCAA
     1741 GTGTATCATA TGCCAAGTAC GCCCCCTATT GACGTCAATG ACGGTAAATG GCCCGCCTGG
     1801 CATTGTGCCC AGTACATGAC CTTATGGGAC TTTCCTACTT GGCAGTACAT CTACGTATTA
     1861 GTCATCGCTA TTACCATGGt cgaggtgagc cccacgttct gcttcactct ccccatctcc
     1921 cccccctccc cacccccaat tttgtattta tttatttttt aattattttg tgcagcgatg
     1981 ggggcggggg gggggggggg gcgcgcgcca ggcggggcgg ggcggggcga ggggcggggc
     2041 ggggcgaggc ggagaggtgc ggcggcagcc aatcagagcg gcgcgctccg aaagtttcct
     2101 tttatggcga ggcggcggcg gcggcggccc tataaaaagc gaagcgcgcg gcgggcggga
     2161 gtcgctgcga cgctgccttc gccccgtgcc ccgctccgcc gccgcctcgc gccgcccgcc
     2221 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
     2281 gggctgtaat tagctgagca agaggtaagg gtttaaggga tggttggttg gtggggtatt
     2341 aatgtttaat tacctggagc acctgcctga aatcactttt tttcaggttg gaccggtgcc
     2401 accatggact ataaggacca cgacggagac tacaaggatc atgatattga ttacaaagac
     2461 gatgacgata agatggcccc aaagaagaag cggaaggtcg gtatccacgg agtcccagca
     2521 gccgacaaga agtacagcat cggcctggac atcggcacca actctgtggg ctgggccgtg
     2581 atcaccgacg agtacaaggt gcccagcaag aaattcaagg tgctgggcaa caccgaccgg
     2641 cacagcatca agaagaacct gatcggagcc ctgctgttcg acagcggcga aacagccgag
     2701 gccacccggc tgaagagaac cgccagaaga agatacacca gacggaagaa ccggatctgc
     2761 tatctgcaag agatcttcag caacgagatg gccaaggtgg acgacagctt cttccacaga
     2821 ctggaagagt ccttcctggt ggaagaggat aagaagcacg agcggcaccc catcttcggc
     2881 aacatcgtgg acgaggtggc ctaccacgag aagtacccca ccatctacca cctgagaaag
     2941 aaactggtgg acagcaccga caaggccgac ctgcggctga tctatctggc cctggcccac
     3001 atgatcaagt tccggggcca cttcctgatc gagggcgacc tgaaccccga caacagcgac
     3061 gtggacaagc tgttcatcca gctggtgcag acctacaacc agctgttcga ggaaaacccc
     3121 atcaacgcca gcggcgtgga cgccaaggcc atcctgtctg ccagactgag caagagcaga
     3181 cggctggaaa atctgatcgc ccagctgccc ggcgagaaga agaatggcct gttcggaaac
     3241 ctgattgccc tgagcctggg cctgaccccc aacttcaaga gcaacttcga cctggccgag
     3301 gatgccaaac tgcagctgag caaggacacc tacgacgacg acctggacaa cctgctggcc
     3361 cagatcggcg accagtacgc cgacctgttt ctggccgcca agaacctgtc cgacgccatc
     3421 ctgctgagcg acatcctgag agtgaacacc gagatcacca aggcccccct gagcgcctct
     3481 atgatcaaga gatacgacga gcaccaccag gacctgaccc tgctgaaagc tctcgtgcgg
     3541 cagcagctgc ctgagaagta caaagagatt ttcttcgacc agagcaagaa cggctacgcc
     3601 ggctacattg acggcggagc cagccaggaa gagttctaca agttcatcaa gcccatcctg
     3661 gaaaagatgg acggcaccga ggaactgctc gtgaagctga acagagagga cctgctgcgg
     3721 aagcagcgga ccttcgacaa cggcagcatc ccccaccaga tccacctggg agagctgcac
     3781 gccattctgc ggcggcagga agatttttac ccattcctga aggacaaccg ggaaaagatc
     3841 gagaagatcc tgaccttccg catcccctac tacgtgggcc ctctggccag gggaaacagc
     3901 agattcgcct ggatgaccag aaagagcgag gaaaccatca ccccctggaa cttcgaggaa
     3961 gtggtggaca agggcgcttc cgcccagagc ttcatcgagc ggatgaccaa cttcgataag
     4021 aacctgccca acgagaaggt gctgcccaag cacagcctgc tgtacgagta cttcaccgtg
     4081 tataacgagc tgaccaaagt gaaatacgtg accgagggaa tgagaaagcc cgccttcctg
     4141 agcggcgagc agaaaaaggc catcgtggac ctgctgttca agaccaaccg gaaagtgacc
     4201 gtgaagcagc tgaaagagga ctacttcaag aaaatcgagt gcttcgactc cgtggaaatc
     4261 tccggcgtgg aagatcggtt caacgcctcc ctgggcacat accacgatct gctgaaaatt
     4321 atcaaggaca aggacttcct ggacaatgag gaaaacgagg acattctgga agatatcgtg
     4381 ctgaccctga cactgtttga ggacagagag atgatcgagg aacggctgaa aacctatgcc
     4441 cacctgttcg acgacaaagt gatgaagcag ctgaagcggc ggagatacac cggctggggc
     4501 aggctgagcc ggaagctgat caacggcatc cgggacaagc agtccggcaa gacaatcctg
     4561 gatttcctga agtccgacgg cttcgccaac agaaacttca tgcagctgat ccacgacgac
     4621 agcctgacct ttaaagagga catccagaaa gcccaggtgt ccggccaggg cgatagcctg
     4681 cacgagcaca ttgccaatct ggccggcagc cccgccatta agaagggcat cctgcagaca
     4741 gtgaaggtgg tggacgagct cgtgaaagtg atgggccggc acaagcccga gaacatcgtg
     4801 atcgaaatgg ccagagagaa ccagaccacc cagaagggac agaagaacag ccgcgagaga
     4861 atgaagcgga tcgaagaggg catcaaagag ctgggcagcc agatcctgaa agaacacccc
     4921 gtggaaaaca cccagctgca gaacgagaag ctgtacctgt actacctgca gaatgggcgg
     4981 gatatgtacg tggaccagga actggacatc aaccggctgt ccgactacga tgtggaccat
     5041 atcgtgcctc agagctttct gaaggacgac tccatcgaca acaaggtgct gaccagaagc
     5101 gacaagaacc ggggcaagag cgacaacgtg ccctccgaag aggtcgtgaa gaagatgaag
     5161 aactactggc ggcagctgct gaacgccaag ctgattaccc agagaaagtt cgacaatctg
     5221 accaaggccg agagaggcgg cctgagcgaa ctggataagg ccggcttcat caagagacag
     5281 ctggtggaaa cccggcagat cacaaagcac gtggcacaga tcctggactc ccggatgaac
     5341 actaagtacg acgagaatga caagctgatc cgggaagtga aagtgatcac cctgaagtcc
     5401 aagctggtgt ccgatttccg gaaggatttc cagttttaca aagtgcgcga gatcaacaac
     5461 taccaccacg cccacgacgc ctacctgaac gccgtcgtgg gaaccgccct gatcaaaaag
     5521 taccctaagc tggaaagcga gttcgtgtac ggcgactaca aggtgtacga cgtgcggaag
     5581 atgatcgcca agagcgagca ggaaatcggc aaggctaccg ccaagtactt cttctacagc
     5641 aacatcatga actttttcaa gaccgagatt accctggcca acggcgagat ccggaagcgg
     5701 cctctgatcg agacaaacgg cgaaaccggg gagatcgtgt gggataaggg ccgggatttt
     5761 gccaccgtgc ggaaagtgct gagcatgccc caagtgaata tcgtgaaaaa gaccgaggtg
     5821 cagacaggcg gcttcagcaa agagtctatc ctgcccaaga ggaacagcga taagctgatc
     5881 gccagaaaga aggactggga ccctaagaag tacggcggct tcgacagccc caccgtggcc
     5941 tattctgtgc tggtggtggc caaagtggaa aagggcaagt ccaagaaact gaagagtgtg
     6001 aaagagctgc tggggatcac catcatggaa agaagcagct tcgagaagaa tcccatcgac
     6061 tttctggaag ccaagggcta caaagaagtg aaaaaggacc tgatcatcaa gctgcctaag
     6121 tactccctgt tcgagctgga aaacggccgg aagagaatgc tggcctctgc cggcgaactg
     6181 cagaagggaa acgaactggc cctgccctcc aaatatgtga acttcctgta cctggccagc
     6241 cactatgaga agctgaaggg ctcccccgag gataatgagc agaaacagct gtttgtggaa
     6301 cagcacaagc actacctgga cgagatcatc gagcagatca gcgagttctc caagagagtg
     6361 atcctggccg acgctaatct ggacaaagtg ctgtccgcct acaacaagca ccgggataag
     6421 cccatcagag agcaggccga gaatatcatc cacctgttta ccctgaccaa tctgggagcc
     6481 cctgccgcct tcaagtactt tgacaccacc atcgaccgga agaggtacac cagcaccaaa
     6541 gaggtgctgg acgccaccct gatccaccag agcatcaccg gcctgtacga gacacggatc
     6601 gacctgtctc agctgggagg cgacaaaagg ccggcggcca cgaaaaaggc cggccaggca
     6661 aaaaagaaaa agcttgaggg cagaggaagt ctgctaacat gcggtgacgt ggaggagaat
     6721 cccggccctg ctagcatggt gagcaagggc gaggaggata acatggccat catcaaggag
     6781 ttcatgcgct tcaaggtgca catggagggc tccgtgaacg gccacgagtt cgagatcgag
     6841 ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa ggtgaccaag
     6901 ggtggccccc tgcccttcgc ctgggacatc ctgtcccctc agttcatgta cggctccaag
     6961 gcctacgtga agcaccccgc cgacatcccc gactacttga agctgtcctt ccccgagggc
     7021 ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg tggtgaccgt gacccaggac
     7081 tcctccctgc aggacggcga gttcatctac aaggtgaagc tgcgcggcac caacttcccc
     7141 tcagacggcc ccgtaatgca gaagaaaacc atgggctggg aggcctcctc cgagcggatg
     7201 taccccgagg acggcgccct gaagggcgag atcaagcaga ggctgaagct gaaggacggc
     7261 ggccactacg acgctgaggt caagaccacc tacaaggcca agaagcccgt gcagctgccc
     7321 ggcgcctaca acgtcaacat caagttggac atcacctccc acaacgagga ctacaccatc
     7381 gtggaacagt acgaacgcgc cgagggccgc cactccaccg gcggcatgga cgagctgtac
     7441 aagtaagaat tcctagagct cgctgatcag cctcgactgt gccttctagt tgccagccat
     7501 ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc
     7561 tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg
     7621 ggggtggggt ggggcaggac agcaaggggg aggattggga agagaatagc aggcatgctg
     7681 gggagcggcc gcaggaaccc ctagtgatgg agttggccac tccctctctg cgcgctcgct
     7741 cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc cgggcggcct
     7801 cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat tttctcctta
     7861 cgcatctgtg cggtatttca caccgcatac gtcaaagcaa ccatagtacg cgccctgtag
     7921 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     7981 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     8041 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     8101 cctcgacccc aaaaaacttg atttgggtga tggttcacgt agtgggccat cgccctgata
     8161 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     8221 aactggaaca acactcaacc ctatctcggg ctattctttt gatttataag ggattttgcc
     8281 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa
     8341 caaaatatta acgtttacaa ttttatggtg cactctcagt acaatctgct ctgatgccgc
     8401 atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct
     8461 gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag
     8521 gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac gcctattttt
     8581 ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa
     8641 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     8701 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     8761 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     8821 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     8881 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     8941 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     9001 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     9061 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     9121 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     9181 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     9241 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     9301 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     9361 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     9421 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtggaagcc gcggtatcat
     9481 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     9541 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     9601 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     9661 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     9721 t
//