LOCUS       Exported                9704 bp ds-DNA     circular SYN 27-OCT-2020
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9704)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 2, 2021 from SnapGene 4.2.11
            http://www.snapgene.com
COMMENT     http://www.informaxinc.com/
FEATURES             Location/Qualifiers
     source          1..9704
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     source          6407..6412
                     /organism="pUC19"
                     /mol_type="other DNA"
                     /label=source:pUC19(3)
     source          6541..6546
                     /organism="pUC19"
                     /mol_type="other DNA"
                     /label=source:pUC19(2)
     source          6669..6674
                     /organism="pUC19"
                     /mol_type="other DNA"
                     /label=source:pUC19(1)
     source          6877..6882
                     /organism="pUC19"
                     /mol_type="other DNA"
                     /label=source:pUC19
     primer_bind     complement(70..87)
                     /label=QB0090 (pQE promoter)
     misc_feature    201..1016
                     /label=Kan
     primer_bind     2142..2164
                     /label=QB0050
     primer_bind     2161..2177
                     /label=M13R (QB0048)
     primer_bind     2188..2207
                     /label=HDR-upF
     primer_bind     complement(3092..3124)
                     /label=HDR-upR
     primer_bind     complement(3114..3141)
                     /label=EZH2HDRup1R
     misc_RNA        complement(3122..3141)
                     /label=sgRNA1
     exon            3132..3141
                     /label=exon1
     misc_feature    3139..3141
                     /label=ATG
     gene            3142..5391
                     /label=Ezh2
     misc_feature    3142..3885
                     /label=N-half of Ezh2
     misc_feature    3232..3357
                     /label=EBD
     misc_feature    3232..3234
                     /label=F32
     misc_feature    3238..3240
                     /label=R34
     misc_feature    3244..3246
                     /label=D36
     misc_feature    3253..3255
                     /label=K39
     misc_feature    3358..3468
                     /label=BAM
     misc_feature    3742..3813
                     /label=202-225
     primer_bind     complement(3745..3765)
                     /label=hsEzh2-seqR
     misc_feature    3766..3768
                     /label=K210
     misc_feature    3787..3789
                     /label=K217
     misc_feature    3802..3804
                     /label=K222
     primer_bind     complement(4602..4623)
                     /label=DY 97 - Ezh2 seq. R
     misc_feature    4660..4683
                     /label=503-510
     misc_feature    4672..4962
                     /label=CXC
     misc_feature    4687..4746
                     /label=512-531
     misc_feature    4963..5391
                     /label=SET
     misc_feature    5098..5130
                     /label=649-659
     primer_bind     complement(5341..5362)
                     /label=xw499
     3'UTR           5395..6266
                     /label=EZH2 3'UTR
     polyA_signal    6285..6406
                     /label=SV40 poly(A) signal(2)
     polyA_signal    6413..6534
                     /label=SV40 poly(A) signal(1)
     polyA_signal    6547..6668
                     /label=SV40 poly(A) signal
     polyA_signal    6674..6877
                     /label=bGH poly(A) signal
     misc_feature    6883..6884
                     /label=Extra Bases
     misc_feature    6885..6918
                     /label=LoxP+2(2)
     misc_feature    6919..6924
                     /label=BamHI
     misc_feature    6925..7193
                     /label=SV40 promoter
     primer_bind     complement(6988..7009)
                     /label=SV40Pro-R (YL114)
     CDS             7259..7855
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     CDS             7865..7918
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             7931..8641
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     misc_feature    8650..8698
                     /label=Synthetic poly(A)
     misc_feature    8699..8704
                     /label=HindIII
     misc_feature    8705..8738
                     /label=LoxP+2
     primer_bind     8744..8771
                     /label=HDR-dnF2
     exon            9282..9410
                     /label=exon2
     primer_bind     complement(9392..9411)
                     /label=HDR-dnR
     primer_bind     complement(9490..9511)
                     /label=HDR-dnR2
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
      121 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
      181 aatattgaaa aaggaagagt atgagccata ttcaacggga aacgtcttgc tctaggccgc
      241 gattaaattc caacatggat gctgatttat atgggtataa atgggctcgc gataatgtcg
      301 ggcaatcagg tgcgacaatc tatcgattgt atgggaagcc cgatgcgcca gagttgtttc
      361 tgaaacatgg caaaggtagc gttgccaatg atgttacaga tgagatggtc agactaaact
      421 ggctgacgga atttatgcct cttccgacca tcaagcattt tatccgtact cctgatgatg
      481 catggttact caccactgcg atccctggga aaacagcatt ccaggtatta gaagaatatc
      541 ctgattcagg tgaaaatatt gttgatgcgc tggcagtgtt cctgcgccgg ttgcattcga
      601 ttcctgtttg taattgtcct tttaacagcg atcgcgtatt tcgtctcgct caggcgcaat
      661 cacgaatgaa taacggtttg gttgatgcga gtgattttga tgacgagcgt aatggctggc
      721 ctgttgaaca agtctggaaa gaaatgcata aacttttgcc attctcaccg gattcagtcg
      781 tcactcatgg tgatttctca cttgataacc ttatttttga cgaggggaaa ttaataggtt
      841 gtattgatgt tggacgagtc ggaatcgcag accgatacca ggatcttgcc atcctatgga
      901 actgcctcgg tgagttttct ccttcattac agaaacggct ttttcaaaaa tatggtattg
      961 ataatcctga tatgaataaa ttgcagtttc atttgatgct cgatgagttt ttctaactgt
     1021 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     1081 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     1141 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     1201 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     1261 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     1321 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     1381 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     1441 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     1501 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     1561 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     1621 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     1681 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     1741 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     1801 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     1861 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     1921 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc
     1981 tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag
     2041 cgggcagtga gcgcaacgca attaatgtga gttagctcac tcattaggca ccccaggctt
     2101 tacactttat gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca
     2161 caggaaacag ctatgaccat gctcgagtca ccgcacaccc ttttacttaa aacactaaaa
     2221 gtggcttctc attgttctta agataaagca caaattgtta gtgtggcctg taaagctttg
     2281 catagcctga cagagaatgt cctgctaata atttgaaggt acaggatgat tttaatactt
     2341 taggagaaaa tgttctagga aaagacgctt gtttagactt aaggtgagga ctctgcagta
     2401 tgaattagac atctggtgaa ctataagctg tccccgcatt taaacataat tggttctgag
     2461 agcctgcaac taaagataag gcagaagaat ttactttgca tttcctgcat tcctcttttc
     2521 gcttgatagc agaaacccct catgttaata aaggtggcac aagaggcaaa aatacagact
     2581 ttatcacagt gtttaaggag aggtgcatga ttaagtgtgt ggggagagag tacctttgta
     2641 cattttatta tatggtgaac tgtatgtttt ctacttttag tactgtttgt aaattttact
     2701 tcttcttgga tttacctttt tcagttatat tattccatta tgccttgcta ctgtaacagc
     2761 taatgatgaa aaacaggatc tgtctttata ttttcttccc tccacaaatg tggatctcat
     2821 agagttgaaa actaggttgt gatatagtat agtataccta attcctgtaa tgggatcatg
     2881 ttcctataat atggccgcaa tttagtgtag aatttttgta aataaaagtg tattttaagt
     2941 ttaacttaaa ctttcaatga agtgttttaa ggatttaacc atgcagcaca aatgagcacc
     3001 tttctgtaaa tgccaacagt gtaatatgtg tcatttcttc actgattgtt agtttgctgc
     3061 ggattaaaac acaggtgatc atattcaggc tggttagatt agtgatttta atatgaaacc
     3121 attgctttta gaataatcat gggccagact gggaagaaat ctgagaaggg accagtttgt
     3181 tggcggaagc gtgtaaaatc agagtacatg cgactgagac agctcaagag gttcagacga
     3241 gctgatgaag taaagagtat gtttagttcc aatcgtcaga aaattttgga aagaacggaa
     3301 atcttaaacc aagaatggaa acagcgaagg atacagcctg tgcacatcct gacttctgtg
     3361 agctcattgc gcgggactag ggagtgttcg gtgaccagtg acttggattt tccaacacaa
     3421 gtcatcccat taaagactct gaatgcagtt gcttcagtac ccataatgta ttcttggtca
     3481 cccctacagc agaattttat ggtggaagat gaaactgttt tacataacat tccttatatg
     3541 ggagatgaag ttttagatca ggatggtact ttcattgaag aactaataaa aaattatgat
     3601 gggaaagtac acggggatag agaatgtggg tttataaatg atgaaatttt tgtggagttg
     3661 gtgaatgccc ttggtcaata taatgatgat gacgatgatg atgatggaga cgatcctgaa
     3721 gaaagagaag aaaagcagaa agatctggag gatcaccgag atgataaaga aagccgccca
     3781 cctcggaaat ttccttctga taaaattttt gaagccattt cctcaatgtt tccagataag
     3841 ggcacagcag aagaactaaa ggaaaaatat aaagaactca ccgaacagca gctcccaggc
     3901 gcacttcctc ctgaatgtac ccccaacata gatggaccaa atgctaaatc tgttcagaga
     3961 gagcaaagct tacactcctt tcatacgctt ttctgtaggc gatgttttaa atatgactgc
     4021 ttcctacatc gtaagtgcaa ttattctttt catgcaacac ccaacactta taagcggaag
     4081 aacacagaaa cagctctaga caacaaacct tgtggaccac agtgttacca gcatttggag
     4141 ggagcaaagg agtttgctgc tgctctcacc gctgagcgga taaagacccc accaaaacgt
     4201 ccaggaggcc gcagaagagg acggcttccc aataacagta gcaggcccag cacccccacc
     4261 attaatgtgc tggaatcaaa ggatacagac agtgataggg aagcagggac tgaaacgggg
     4321 ggagagaaca atgataaaga agaagaagag aagaaagatg aaacttcgag ctcctctgaa
     4381 gcaaattctc ggtgtcaaac accaataaag atgaagccaa atattgaacc tcctgagaat
     4441 gtggagtgga gtggtgctga agcctcaatg tttagagtcc tcattggcac ttactatgac
     4501 aatttctgtg ccattgctag gttaattggg accaaaacat gtagacaggt gtatgagttt
     4561 agagtcaaag aatctagcat catagctcca gctcccgctg aggatgtgga tactcctcca
     4621 aggaaaaaga agaggaaaca ccggttgtgg gctgcacact gcagaaagat acagctgaaa
     4681 aaggacggct cctctaacca tgtttacaac tatcaaccct gtgatcatcc acggcagcct
     4741 tgtgacagtt cgtgcccttg tgtgatagca caaaattttt gtgaaaagtt ttgtcaatgt
     4801 agttcagagt gtcaaaaccg ctttccggga tgccgctgca aagcacagtg caacaccaag
     4861 cagtgcccgt gctacctggc tgtccgagag tgtgaccctg acctctgtct tacttgtgga
     4921 gccgctgacc attgggacag taaaaatgtg tcctgcaaga actgcagtat tcagcggggc
     4981 tccaaaaagc atctattgct ggcaccatct gacgtggcag gctgggggat ttttatcaaa
     5041 gatcctgtgc agaaaaatga attcatctca gaatactgtg gagagattat ttctcaagat
     5101 gaagctgaca gaagagggaa agtgtatgat aaatacatgt gcagctttct gttcaacttg
     5161 aacaatgatt ttgtggtgga tgcaacccgc aagggtaaca aaattcgttt tgcaaatcat
     5221 tcggtaaatc caaactgcta tgcaaaagtt atgatggtta acggtgatca caggataggt
     5281 atttttgcca agagagccat ccagactggc gaagagctgt tttttgatta cagatacagc
     5341 caggctgatg ccctgaagta tgtcggcatc gaaagagaaa tggaaatccc ttgacatctg
     5401 ctacctcctc ccccctcctc tgaaacagct gccttagctt caggaacctc gagtactgtg
     5461 ggcaatttag aaaaagaaca tgcagtttga aattctgaat ttgcaaagta ctgtaagaat
     5521 aatttatagt aatgagttta aaaatcaact ttttattgcc ttctcaccag ctgcaaagtg
     5581 ttttgtacca gtgaattttt gcaataatgc agtatggtac atttttcaac tttgaataaa
     5641 gaatacttga acttgtcctt gttgaatcat ctctcataac gtgtcaataa ctgctttcag
     5701 gactgctcaa caaaaaactt ttaaatttaa ctccttgggt tttttagttg tctttgtatg
     5761 aatcttgatt taactaatac tgcctgtgaa tatgtggaag acatatatga agaacactgc
     5821 tgtgcatcct gagggcttac ccagtaaagg gcttgatctt atagacaaga ccagggtggg
     5881 tacgttccag tcagctgtgc aacctggaga aagagattta acctccaagt ctgttcccca
     5941 tatataaact gacagagtag acttttaggt tatttttaag aacgtaccta gttctaaact
     6001 ccattagtct agatagagtt ctttagaaac aaacacaata taagttggga ttccttacct
     6061 accttatcac aaaaggtgat tgataggtat aaccaaactg tagttttata tcttctcaga
     6121 ggtaaaaatc acaaactctg aaagcgtcaa ggaaacgaca tgtaccttgc atgccttatg
     6181 ataatgtggt ttgtcccaga atttgaggta acttccctgc accctcccca ggtacattca
     6241 ccagcagcat catgggaacc ctgacaacta gtagatctgg taccaacttg tttattgcag
     6301 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     6361 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttactgc agaacttgtt
     6421 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     6481 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttactgcag
     6541 gtcgacaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     6601 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     6661 gtatcttagg atccgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc
     6721 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa
     6781 attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac
     6841 agcaaggggg aggattggga agagaatagc aggcatggta ccggataact tcgtatagca
     6901 tacattatac gaagttatgg atccggtgtg gaaagtcccc aggctcccca gcaggcagaa
     6961 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     7021 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     7081 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     7141 gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga
     7201 agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcgct agcccaccat
     7261 gaccgagtac aagcccacgg tgcgcctcgc cacccgcgac gacgtccccc gggccgtacg
     7321 caccctcgcc gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg acccggaccg
     7381 ccacatcgag cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat
     7441 cggcaaggtg tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag
     7501 cgtcgaagcg ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc
     7561 ccggctggcc gcgcagcaac agatggaagg cctcctggcg ccgcaccggc ccaaggagcc
     7621 cgcgtggttc ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag
     7681 cgccgtcgtg ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga
     7741 gacctccgcg ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga
     7801 cgtcgaggtg cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgccggatc
     7861 gggagagggc agaggaagtc tgctaacatg cggtgacgtc gaggagaatc ctggcccacc
     7921 ggtcgccacc atggtgagca agggcgagga ggataacatg gccatcatca aggagttcat
     7981 gcgcttcaag gtgcacatgg agggctccgt gaacggccac gagttcgaga tcgagggcga
     8041 gggcgagggc cgcccctacg agggcaccca gaccgccaag ctgaaggtga ccaagggtgg
     8101 ccccctgccc ttcgcctggg acatcctgtc ccctcagttc atgtacggct ccaaggccta
     8161 cgtgaagcac cccgccgaca tccccgacta cttgaagctg tccttccccg agggcttcaa
     8221 gtgggagcgc gtgatgaact tcgaggacgg cggcgtggtg accgtgaccc aggactcctc
     8281 cctgcaggac ggcgagttca tctacaaggt gaagctgcgc ggcaccaact tcccctccga
     8341 cggccccgta atgcagaaga agaccatggg ctgggaggcc tcctccgagc ggatgtaccc
     8401 cgaggacggc gccctgaagg gcgagatcaa gcagaggctg aagctgaagg acggcggcca
     8461 ctacgacgct gaggtcaaga ccacctacaa ggccaagaag cccgtgcagc tgcccggcgc
     8521 ctacaacgtc aacatcaagt tggacatcac ctcccacaac gaggactaca ccatcgtgga
     8581 acagtacgaa cgcgccgagg gccgccactc caccggcggc atggacgagc tgtacaagta
     8641 agcggccgca ataaaatatc tttattttca ttacatctgt gtgttggttt tttgtgtgaa
     8701 gcttataact tcgtatagca tacattatac gaagttatct agagtttttt cccctcctaa
     8761 gttcaataag tttttattta aaataattct agccaagttt gtatttacaa taagatgagt
     8821 gaatacttgg aagcttaggt tcttgatctg tcttaacatt ataccatgca cattaatcag
     8881 tctttgttta gtatactttt atttgcatat gcagctgcag aaatgcctcc ctaaatataa
     8941 agggatgcat ttcatgattt tatgtttcta aagtcattgt gattaaaaaa aaaaatacac
     9001 acacacacac acacacacac acacacacac acacacacac agtcaacaaa gaaaggatgg
     9061 caatcgtttc ctgttctcta accacttgca ataaaataac agatgtggtg tttttgtatt
     9121 atttgaatgt gggaaactat agtgtaaaat ctatatgcac ctaagaaaat gttcttgata
     9181 tttttgagga tacactagat ttttaacctg ctttacaggt gttatacatt ttctaacaat
     9241 gaacaatttc tcctttcctc tccttcattt ttttttttaa gagtatgttt agttccaatc
     9301 gtcagaaaat tttggaaaga acggaaatct taaaccaaga atggaaacag cgaaggatac
     9361 agcctgtgca catcctgact tctgtgagct cattgcgcgg gactagggag gttggttaac
     9421 ataacatgtt gtagtaatta tcatcatctt ttggtttatg ctattggaac atttgtttgg
     9481 ttattgggag gaaatcattg acctcagggt ggaattcata tggtgcactc tcagtacaat
     9541 ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc
     9601 ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag
     9661 ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcga
//
