LOCUS       SaCas9v2_Puro_(copy)    8083 bp ds-DNA     circular     05-NOV-2021
DEFINITION  .
FEATURES             Location/Qualifiers
     misc_feature    1..241
                     /label="hU6 promoter"
                     /ApEinfo_revcolor=#b4abac
                     /ApEinfo_fwdcolor=#b4abac
     primer_bind     172..191
                     /label="LKO.1 5'"
                     /ApEinfo_revcolor=#b7e6d7
                     /ApEinfo_fwdcolor=#b7e6d7
     misc_RNA        272..343
                     /label="Sa gRNA scaffold"
                     /ApEinfo_revcolor=#c6c9d1
                     /ApEinfo_fwdcolor=#c6c9d1
     enhancer        363..648
                     /label="CMV enhancer"
                     /ApEinfo_revcolor=#b1ff67
                     /ApEinfo_fwdcolor=#b1ff67
     promoter        650..927
                     /label="chicken beta-actin promoter"
                     /ApEinfo_revcolor=#b7e6d7
                     /ApEinfo_fwdcolor=#b7e6d7
     intron          928..1155
                     /label="hybrid intron"
                     /ApEinfo_revcolor=#faac61
                     /ApEinfo_fwdcolor=#faac61
     regulatory      1167..1176
                     /label="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)"
                     /ApEinfo_revcolor=#c6c9d1
                     /ApEinfo_fwdcolor=#c6c9d1
     CDS             1173..5096
                     /label="Translation 1173-5096"
     CDS             1179..1199
                     /label="SV40 NLS"
                     /ApEinfo_revcolor=#b4abac
                     /ApEinfo_fwdcolor=#b4abac
     CDS             1224..4379
                     /label="SaCas9"
                     /ApEinfo_revcolor=#b7e6d7
                     /ApEinfo_fwdcolor=#b7e6d7
     misc_feature    4380..4427
                     /label="Nucleoplasmin"
                     /ApEinfo_revcolor=#f58a5e
                     /ApEinfo_fwdcolor=#f58a5e
     CDS             4443..4496
                     /label="T2A"
                     /ApEinfo_revcolor=#c6c9d1
                     /ApEinfo_fwdcolor=#c6c9d1
     primer_bind     complement(4497..4516)
                     /label="Puro-R"
                     /ApEinfo_revcolor=#c7b0e3
                     /ApEinfo_fwdcolor=#c7b0e3
     CDS             4497..5096
                     /label="PuroR"
                     /ApEinfo_revcolor=#9eafd2
                     /ApEinfo_fwdcolor=#9eafd2
     primer_bind     4993..5013
                     /label="Puro-F"
                     /ApEinfo_revcolor=#ff9ccd
                     /ApEinfo_fwdcolor=#ff9ccd
     primer_bind     complement(5124..5141)
                     /label="BGH-rev"
                     /ApEinfo_revcolor=#ffef86
                     /ApEinfo_fwdcolor=#ffef86
     polyA_signal    5130..5337
                     /label="bGH poly(A) signal"
                     /ApEinfo_revcolor=#b4abac
                     /ApEinfo_fwdcolor=#b4abac
     repeat_region   5346..5475
                     /label="AAV2 ITR"
                     /ApEinfo_revcolor=#faac61
                     /ApEinfo_fwdcolor=#faac61
     repeat_region   5346..5486
                     /label="AAV2 ITR"
                     /ApEinfo_revcolor=#9eafd2
                     /ApEinfo_fwdcolor=#9eafd2
     rep_origin      5561..6016
                     /label="f1 ori"
                     /ApEinfo_revcolor=#d6b295
                     /ApEinfo_fwdcolor=#d6b295
     primer_bind     complement(5648..5667)
                     /label="F1ori-R"
                     /ApEinfo_revcolor=#faac61
                     /ApEinfo_fwdcolor=#faac61
     primer_bind     5858..5879
                     /label="F1ori-F"
                     /ApEinfo_revcolor=#c6c9d1
                     /ApEinfo_fwdcolor=#c6c9d1
     primer_bind     complement(6033..6052)
                     /label="pRS-marker"
                     /ApEinfo_revcolor=#ffef86
                     /ApEinfo_fwdcolor=#ffef86
     primer_bind     6152..6174
                     /label="pGEX 3'"
                     /ApEinfo_revcolor=#d59687
                     /ApEinfo_fwdcolor=#d59687
     primer_bind     complement(6212..6230)
                     /label="pBRforEco"
                     /ApEinfo_revcolor=#c6c9d1
                     /ApEinfo_fwdcolor=#c6c9d1
     promoter        6298..6402
                     /label="AmpR promoter"
                     /ApEinfo_revcolor=#c7b0e3
                     /ApEinfo_fwdcolor=#c7b0e3
     CDS             6403..7263
                     /label="AmpR"
                     /ApEinfo_revcolor=#84b0dc
                     /ApEinfo_fwdcolor=#84b0dc
     primer_bind     complement(6621..6640)
                     /label="Amp-R"
                     /ApEinfo_revcolor=#75c6a9
                     /ApEinfo_fwdcolor=#75c6a9
     rep_origin      7434..8022
                     /label="ori"
                     /ApEinfo_revcolor=#d59687
                     /ApEinfo_fwdcolor=#d59687
     primer_bind     7923..7942
                     /label="pBR322ori-F"
                     /ApEinfo_revcolor=#b7e6d7
                     /ApEinfo_fwdcolor=#b7e6d7
ORIGIN
        1 GAGGGCCTAT TTCCCATGAT TCCttcatat ttgcatatac gatacaaggc tgttagagag
       61 ataattggaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
      121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
      181 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
      241 cgaaacaccg ggtcttcgag aagacctgtt ttagtactct ggaaacagaa tctactaaaa
      301 caaggcaaaa tgccgtgttt atctcgtcaa cttgttggcg agaTTTTTGC GGCCGCggta
      361 cccgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca
      421 ttgacgtcaa tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta
      481 cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt
      541 gacgtcaatg acggtaaatg gcccgcctgg cattgtgccc agtacatgac cttatgggac
      601 tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt cgaggtgagc
      661 cccacgttct gcttcactct ccccatctcc cccccctccc cacccccaat tttgtattta
      721 tttatttttt aattattttg tgcagcgatg ggggcggggg gggggggggg gcgcgcgcca
      781 ggcggggcgg ggcggggcga ggggcggggc ggggcgaggc ggagaggtgc ggcggcagcc
      841 aatcagagcg gcgcgctccg aaagtttcct tttatggcga ggcggcggcg gcggcggccc
      901 tataaaaagc gaagcgcgcg gcgggcggga gtcgctgcgc gctgccttcg ccccgtgccc
      961 cgctccgccg ccgcctcgcg ccgcccgccc cggctctgac tgaccgcgtt actcccacag
     1021 gtgagcgggc gggacggccc ttctcctccg ggctgtaatt agctgagcaa gaggtaaggg
     1081 tttaagggat ggttggttgg tggggtatta atgtttaatt acctggagca cctgcctgaa
     1141 atcacttttt ttcaggttgg accggtgcca ccatggcccc aaagaagaag cggaaggtcg
     1201 gtatccacgg agtcccagca gccaagcgga actacatcct gggcctggac atcggcatca
     1261 ccagcgtggg ctacggcatc atcgactacg agacacggga cgtgatcgat gccggcgtgc
     1321 ggctgttcaa agaggccaac gtggaaaaca acgagggcag gcggagcaag agaggcgcca
     1381 gaaggctgaa gcggcggagg cggcatagaa tccagagagt gaagaagctg ctgttcgact
     1441 acaacctgct gaccgaccac agcgagctga gcggcatcaa cccctacgag gccagagtga
     1501 agggcctgag ccagaagctg agcgaggaag agttctctgc cgccctgctg cacctggcca
     1561 agagaagagg cgtgcacaac gtgaacgagg tggaagagga caccggcaac gagctgtcca
     1621 ccaaagagca gatcagccgg aacagcaagg ccctggaaga gaaatacgtg gccgaactgc
     1681 agctggaacg gctgaagaaa gacggcgaag tgcggggcag catcaacaga ttcaagacca
     1741 gcgactacgt gaaagaagcc aaacagctgc tgaaggtgca gaaggcctac caccagctgg
     1801 accagagctt catcgacacc tacatcgacc tgctggaaac ccggcggacc tactatgagg
     1861 gacctggcga gggcagcccc ttcggctgga aggacatcaa agaatggtac gagatgctga
     1921 tgggccactg cacctacttc cccgaggaac tgcggagcgt gaagtacgcc tacaacgccg
     1981 acctgtacaa cgccctgaac gacctgaaca atctcgtgat caccagggac gagaacgaga
     2041 agctggaata ttacgagaag ttccagatca tcgagaacgt gttcaagcag aagaagaagc
     2101 ccaccctgaa gcagatcgcc aaagaaatcc tcgtgaacga agaggatatt aagggctaca
     2161 gagtgaccag caccggcaag cccgagttca ccaacctgaa ggtgtaccac gacatcaagg
     2221 acattaccgc ccggaaagag attattgaga acgccgagct gctggatcag attgccaaga
     2281 tcctgaccat ctaccagagc agcgaggaca tccaggaaga actgaccaat ctgaactccg
     2341 agctgaccca ggaagagatc gagcagatct ctaatctgaa gggctatacc ggcacccaca
     2401 acctgagcct gaaggccatc aacctgatcc tggacgagct gtggcacacc aacgacaacc
     2461 agatcgctat cttcaaccgg ctgaagctgg tgcccaagaa ggtggacctg tcccagcaga
     2521 aagagatccc caccaccctg gtggacgact tcatcctgag ccccgtcgtg aagagaagct
     2581 tcatccagag catcaaagtg atcaacgcca tcatcaagaa gtacggcctg cccaacgaca
     2641 tcattatcga gctggcccgc gagaagaact ccaaggacgc ccagaaaatg atcaacgaga
     2701 tgcagaagcg gaaccggcag accaacgagc ggatcgagga aatcatccgg accaccggca
     2761 aagagaacgc caagtacctg atcgagaaga tcaagctgca cgacatgcag gaaggcaagt
     2821 gcctgtacag cctggaagcc atccctctgg aagatctgct gaacaacccc ttcaactatg
     2881 aggtggacca catcatcccc agaagcgtgt ccttcgacaa cagcttcaac aacaaggtgc
     2941 tcgtgaagca ggaagaaaac agcaagaagg gcaaccggac cccattccag tacctgagca
     3001 gcagcgacag caagatcagc tacgaaacct tcaagaagca catcctgaat ctggccaagg
     3061 gcaagggcag aatcagcaag accaagaaag agtatctgct ggaagaacgg gacatcaaca
     3121 ggttctccgt gcagaaagac ttcatcaacc ggaacctggt ggataccaga tacgccacca
     3181 gaggcctgat gaacctgctg cggagctact tcagagtgaa caacctggac gtgaaagtga
     3241 agtccatcaa tggcggcttc accagctttc tgcggcggaa gtggaagttt aagaaagagc
     3301 ggaacaaggg gtacaagcac cacgccgagg acgccctgat cattgccaac gccgatttca
     3361 tcttcaaaga gtggaagaaa ctggacaagg ccaaaaaagt gatggaaaac cagatgttcg
     3421 aggaaaagca ggccgagagc atgcccgaga tcgaaaccga gcaggagtac aaagagatct
     3481 tcatcacccc ccaccagatc aagcacatta aggacttcaa ggactacaag tacagccacc
     3541 gggtggacaa gaagcctaat agagagctga ttaacgacac cctgtactcc acccggaagg
     3601 acgacaaggg caacaccctg atcgtgaaca atctgaacgg cctgtacgac aaggacaatg
     3661 acaagctgaa aaagctgatc aacaagagcc ccgaaaagct gctgatgtac caccacgacc
     3721 cccagaccta ccagaaactg aagctgatta tggaacagta cggcgacgag aagaatcccc
     3781 tgtacaagta ctacgaggaa accgggaact acctgaccaa gtactccaaa aaggacaacg
     3841 gccccgtgat caagaagatt aagtattacg gcaacaaact gaacgcccat ctggacatca
     3901 ccgacgacta ccccaacagc agaaacaagg tcgtgaagct gtccctgaag ccctacagat
     3961 tcgacgtgta cctggacaat ggcgtgtaca agttcgtgac cgtgaagaat ctggatgtga
     4021 tcaaaaaaga aaactactac gaagtgaata gcaagtgcta tgaggaagct aagaagctga
     4081 agaagatcag caaccaggcc gagtttatcg cctccttcta caacaacgat ctgatcaaga
     4141 tcaacggcga gctgtataga gtgatcggcg tgaacaacga cctgctgaac cggatcgaag
     4201 tgaacatgat cgacatcacc taccgcgagt acctggaaaa catgaacgac aagaggcccc
     4261 ccaggatcat taagacaatc gcctccaaga cccagagcat taagaagtac agcacagaca
     4321 ttctgggcaa cctgtatgaa gtgaaatcta agaagcaccc tcagatcatc aaaaagggca
     4381 aaaggccggc ggccacgaaa aaggccggcc aggcaaaaaa gaaaaaggaa ttcggcagtg
     4441 gagagggcag aggaagtctg ctaacatgcg gtgacgtcga ggagaatcct ggcccaatga
     4501 ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga cgtccccagg gccgtacgca
     4561 ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca caccgtcgat ccggaccgcc
     4621 acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg
     4681 gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt ctggaccacg ccggagagcg
     4741 tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat ggccgagttg agcggttccc
     4801 ggctggccgc gcagcaacag atggaaggcc tcctggcgcc gcaccggccc aaggagcccg
     4861 cgtggttcct ggccaccgtc ggagtctcgc ccgaccacca gggcaagggt ctgggcagcg
     4921 ccgtcgtgct ccccggagtg gaggcggccg agcgcgccgg ggtgcccgcc ttcctggaga
     4981 cctccgcgcc ccgcaacctc cccttctacg agcggctcgg cttcaccgtc accgccgacg
     5041 tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg caagcccggt gcctgagaat
     5101 tctaactaga gctcgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt
     5161 ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta
     5221 ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg
     5281 ggtggggcag gacagcaagg gggaggattg ggaagagaat agcaggcatg ctggggagcg
     5341 gccgcaggaa cccctagtga tggagttggc cactccctct ctgcgcgctc gctcgctcac
     5401 tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag
     5461 cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg tattttctcc ttacgcatct
     5521 gtgcggtatt tcacaccgca tacgtcaaag caaccatagt acgcgccctg tagcggcgca
     5581 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcctta
     5641 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     5701 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     5761 cccaaaaaac ttgatttggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     5821 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     5881 acaacactca actctatctc gggctattct tttgatttat aagggatttt gccgatttcg
     5941 gtctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata
     6001 ttaacgttta caattttatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     6061 agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg
     6121 gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca
     6181 ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt
     6241 aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc
     6301 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     6361 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     6421 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     6481 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     6541 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     6601 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     6661 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     6721 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     6781 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     6841 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     6901 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     6961 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     7021 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     7081 tggtttattg ctgataaatc tggagccggt gagcgtggaa gccgcggtat cattgcagca
     7141 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     7201 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     7261 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     7321 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     7381 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     7441 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     7501 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     7561 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     7621 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     7681 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     7741 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     7801 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     7861 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     7921 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     7981 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     8041 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgt
//