LOCUS       Exported                7360 bp ds-DNA     circular SYN 31-OCT-2019
DEFINITION  Xm-0209.
ACCESSION   .
VERSION     .
KEYWORDS    pCAG_Xph20-SNAPf-CCR5TC
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7360)
  AUTHORS   Matthieu Sainlos
  TITLE     Direct Submission
  JOURNAL   Exported Thursday, Oct 31, 2019 from SnapGene 5.0.4
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7360
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     protein_bind    24..35
                     /label=ZF binding site
                     /bound_moiety="CCR5TC zinc-finger domain"
                     /note="target sequence for the CCR5TC zinc-finger domain 
                     from the CCR5-224 (+) zinc-finger nuclease (Perez et al., 
                     2008; Gross et al., 2013)"
     enhancer        39..418
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        421..696
                     /label=chicken beta-actin promoter
     intron          697..1708
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1716..1735
                     /label=pCAG-F
                     /note="From Addgene list"
     regulatory      1780..1789
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             1795..2079
                     /codon_start=1
                     /product="Paralog-specific PSD-95 binder Xph20 (Rimbault et
                     al., 2019)"
                     /label=Xph20
                     /translation="SSVSSVPTKLEVVAATPTSLLISWDAVAKNVKVGYYRITYGETGG
                     NSPVQEFTVPGSSSTATISGLSPGVDYTITVYANGVLSKMVLPISINYRT"
     CDS             2110..2652
                     /codon_start=1
                     /product="human O6-alkylguanine-DNA-alkyltransferase 
                     variant that forms covalent bonds with benzylguanine 
                     derivatives"
                     /label=SNAP-tag(R)
                     /note="fast-reacting variant, optimized for mammalian 
                     expression"
                     /translation="DKDCEMKRTTLDSPLGKLELSGCEQGLHRIIFLGKGTSAADAVEV
                     PAPAAVLGGPEPLMQATAWLNAYFHQPEAIEEFPVPALHHPVFQQESFTRQVLWKLLKV
                     VKFGEVISYSHLAALAGNPAATAAVKTALSGNPVPILIPCHRVVQGDLDVGGYEGGLAV
                     KEWLLAHEGHRLGKPGLG"
     CDS             2722..3063
                     /codon_start=1
                     /product="zinc-finger domain from the CCR5-224 (+) 
                     zinc-finger nuclease that recognizes the left half-site 
                     target sequence in the human CCR5 gene (Perez et al., 2008;
                     Gross et al., 2013)"
                     /label=CCR5TC
                     /translation="FQCRICMRNFSDRSNLSRHIRTHTGEKPFACDICGRKFAISSNLN
                     SHTKIHTGSQKPFQCRICMRNFSRSDNLARHIRTHTGEKPFACDICGRKFATSGNLTRH
                     TKIHLRGSQL"
     primer_bind     complement(2768..2787)
                     /label=Xos-0059 pCAG-rev
     CDS             3112..3237
                     /codon_start=1
                     /product="Kruppel-associated box (KRAB) transcriptional 
                     repression domain from the rat zinc finger protein Kid-1 
                     (Witzgall et al., 1994)"
                     /label=KRAB-A
                     /translation="VSVTFEDVAVLFTRDEWKKLDLSQRSLYREVMLENYSNLASM"
     intron          3286..3418
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     misc_feature    3427..4015
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3480..3500)
                     /label=WPRE-R
                     /note="From Addgene"
     CDS             complement(3898..3909)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4048..4159
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     polyA_signal    4451..4505
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4864..4880)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    4888..4904
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(4912..4942)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4957..4978
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5037..5232
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5083..5218
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5238..5372
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(5611..6199)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(6668..7045)
                     /label=ISS
                     /note="immunostimulatory sequence from the AmpR gene; 
                     contains unmethylated CpG dinucleotides in the context of 
                     5'-AACGTT-3' (Sato et al., 1996)"
     promoter        complement(7230..7334)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agagtcatcc tcatcgtcga cattgattat tgactagtta
       61 ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac
      121 ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc
      181 aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt
      241 ggactattta cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac
      301 gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac
      361 cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggg
      421 tcgaggtgag ccccacgttc tgcttcactc tccccatctc ccccccctcc ccacccccaa
      481 ttttgtattt atttattttt taattatttt gtgcagcgat gggggcgggg gggggggggg
      541 cgcgcgccag gcggggcggg gcgggggcga gggcggggcg gggcgaggcg gagaggtgcg
      601 gcggcagcca atcagagcgg cgcgctccga aagtttcctt ttatggcgag gcggcggcgg
      661 cggcggccct ataaaaagcg aagcgcgcgg cgggcgggag tcgctgcgtt gccttcgccc
      721 cgtgccccgc tccgcgccgc ctcgcgccgc ccgccccggc tctgactgac cgcgttactc
      781 ccacaggtga gcgggcggga cggcccttct cctccgggct gtaattagcg cttggtttaa
      841 tgacggctcg tttcttttct gtggctgcgt gaaagcctta aagggctccg ggagggccct
      901 ttgtgcgggg gggagcggct cggggggtgc gtgcgtgtgt gtgtgcgtgg ggagcgccgc
      961 gtgcggcccg cgctgcccgg cggctgtgag cgctgcgggc gcggcgcggg gctttgtgcg
     1021 ctccgcgtgt gcgcgagggg agcgcggccg ggggcggtgc cccgcggtgc gggggggctg
     1081 cgaggggaac aaaggctgcg tgcggggtgt gtgcgtgggg gggtgagcag ggggtgtggg
     1141 cgcggcggtc gggctgtaac ccccccctgc acccccctcc ccgagttgct gagcacggcc
     1201 cggcttcggg tgcggggctc cgtgcggggc gtggcgcggg gctcgccgtg ccgggcgggg
     1261 ggtggcggca ggtgggggtg ccgggcgggg cggggccgcc tcgggccggg gagggctcgg
     1321 gggaggggcg cggcggcccc ggagcgccgg cggctgtcga ggcgcggcga gccgcagcca
     1381 ttgcctttta tggtaatcgt gcgagagggc gcagggactt cctttgtccc aaatctgtgc
     1441 ggagccgaaa tctgggaggc gccgccgcac cccctctagc gggcgcgggg cgaagcggtg
     1501 cggcgccggc aggaaggaaa tgggcgggga gggccttcgt gcgtcgccgc gccgccgtcc
     1561 ccttctccct ctccagcctc ggggctgtcc gcggggggac ggctgccttc gggggggacg
     1621 gggcagggcg gggttcggct tctggcgtgt gaccggcggc tctagagcct ctgctaacca
     1681 tgttcatgcc ttcttctttt tcctacagct cctgggcaac gtgctggtta ttgtgctgtc
     1741 tcatcatttt ggcaaagaat tcggtaccgc gggcccgggg ccaccatggc gggcagctct
     1801 gtcagttccg tgccgaccaa actggaagtg gttgcggcca ccccgacgtc tctgctgatc
     1861 agctgggatg cagtggcgaa gaatgtgaag gtcggttatt accgtatcac ctatggcgaa
     1921 acgggcggta actccccggt tcaggaattc accgtcccgg gtagttcctc aaccgcaacg
     1981 atttccggcc tgtcaccggg tgtcgactat accattacgg tttacgctaa tggggtgctg
     2041 agtaagatgg tgcttccgat ttctatcaat taccgtaccg gtagcggctc cagtagatct
     2101 ggtagcggcg acaaagactg cgaaatgaag cgcaccaccc tggatagccc tctgggcaag
     2161 ctggaactgt ctgggtgcga acagggcctg caccgtatca tcttcctggg caaaggaaca
     2221 tctgccgccg acgccgtgga agtgcctgcc ccagccgccg tgctgggcgg accagagcca
     2281 ctgatgcagg ccaccgcctg gctcaacgcc tactttcacc agcctgaggc catcgaggag
     2341 ttccctgtgc cagccctgca ccacccagtg ttccagcagg agagctttac ccgccaggtg
     2401 ctgtggaaac tgctgaaagt ggtgaagttc ggagaggtca tcagctacag ccacctggcc
     2461 gccctggccg gcaatcccgc cgccaccgcc gccgtgaaaa ccgccctgag cggaaatccc
     2521 gtgcccattc tgatcccctg ccaccgggtg gtgcagggcg acctggacgt ggggggctac
     2581 gagggcgggc tcgccgtgaa agagtggctg ctggcccacg agggccacag actgggcaag
     2641 cctgggctgg gtcctgcagg cggtagcggc tcctctgcta gcggagctgg cgctggagcg
     2701 ggtgcagggg ctggctctag attccagtgc cggatctgca tgcggaactt cagcgaccgg
     2761 tccaacctga gcaggcacat cagaacccac accggagaaa agcccttcgc ctgcgacatt
     2821 tgcggccgga agttcgccat cagcagcaac ctgaacagcc acaccaagat ccacactggc
     2881 agccagaaac ctttccagtg cagaatttgt atgagaaact ttagcagaag cgacaacctg
     2941 gccagacaca tccggacaca tactggtgaa aaaccttttg cctgtgatat ctgtggcaga
     3001 aagtttgcca cctccggcaa tctgacccgg cacacaaaga ttcacctgcg gggcagccag
     3061 ctatcgattg tcgacgctcc tgaacaacgt gaaggtgctt ctcaagtttc tgtttctgtt
     3121 acttttgaag atgttgctgt tctttttact cgtgatgaat ggaaaaaact tgatctttct
     3181 caacgttctc tttatcgtga agttatgctt gaaaattatt ctaatcttgc ttctatggct
     3241 taaacgcgtt gtacaagtaa agcggccccg gactcgaggc cgcaggtaag tatcaaggtt
     3301 acaagacagg tttaaggaga ccaatagaaa ctgggcttgt cgagacagag aagactcttg
     3361 cgtttctgat aggcacctat tggtcttact gacatccact ttgcctttct ctccacaggt
     3421 gtcgacaatc aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat
     3481 gttgctcctt ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct
     3541 tcccgtatgg ctttcatttt ctcctccttg tataaatcct ggttgctgtc tctttatgag
     3601 gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc
     3661 cccactggtt ggggcattgc caccacctgt cagctccttt ccgggacttt cgctttcccc
     3721 ctccctattg ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct
     3781 cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga agctgacgtc ctttccatgg
     3841 ctgctcgcct gtgttgccac ctggattctg cgcgggacgt ccttctgcta cgtcccttcg
     3901 gccctcaatc cagcggacct tccttcccgc ggcctgctgc cggctctgcg gcctcttccg
     3961 cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt gggccgcctc cccgcctgga
     4021 attcgagctc ggtacgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt
     4081 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
     4141 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
     4201 tggggcagga cagcaagggg gaggattggg aagacaatag caggccagct tttgttccct
     4261 ttagggttaa ttgcgcgctt ggcgctaatc atggtcatag ctgtttccac tcctcaggtg
     4321 caggctgcct atcagaaggt ggtggctggt gtggccaatg ccctggctca caaataccac
     4381 tgagatcgtt ttccctctgc caaaaattat ggggacatca tgaagcccct tgagcatctg
     4441 acttctggct aataaaggaa atttattttc attgcaatag tgtgttgaat tttttgtgtc
     4501 tctcactcgg aaggacatat gggagggcaa atcatttaaa acatcagaat gagtatttgg
     4561 tttagagttt ggcaacatat gccatatgct ggctgccatg aacaaaggtg gctataaaga
     4621 ggtcatcagt atatgaaaca gccccctgct gtccattcct tattccatag aaaagccttg
     4681 acttgaggtt agattttttt ttatattttg tttgtgttat ttttttcttt aacatcccta
     4741 aaattttcct tacatgtttt actagccaga tttttcctcc tctcctgact actcccagtc
     4801 atagctgtcc ctcttctctt atgaagatcc ctcgacctgc agcccaagct tggcgtaatc
     4861 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     4921 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     4981 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc ggatccgcat
     5041 ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg
     5101 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc
     5161 gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt tggaggccta
     5221 ggcttttgca aaaagctaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     5281 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     5341 aactcatcaa tgtatcttat catgtctgga tccgctgcat taatgaatcg gccaacgcgc
     5401 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     5461 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     5521 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     5581 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     5641 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     5701 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     5761 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcaatgct cacgctgtag
     5821 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     5881 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     5941 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     6001 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt
     6061 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     6121 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     6181 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     6241 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     6301 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     6361 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     6421 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     6481 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     6541 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     6601 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     6661 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     6721 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     6781 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     6841 gttatcactc atggttatgg agcactgcat aattctctta ctgtcatgcc atccgtaaga
     6901 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     6961 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     7021 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     7081 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     7141 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     7201 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     7261 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     7321 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
//
