LOCUS       sequence-341954-        9779 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of NLS-RfxCas13d-NLS-HA .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9779)
  AUTHORS   Mendez-Mancilla A, Wessels HH, Legut M, Kadina A, Mabuchi M, Walker 
            J, Robb GB, Holden K, Sanjana NE
  TITLE     Chemically modified guide RNAs enhance CRISPR-Cas13 knockdown in 
            human cells.
  JOURNAL   Cell Chem Biol. 2022 Feb 17;29(2):321-327.e4. doi: 
            10.1016/j.chembiol.2021.07.011. Epub 2021 Aug 2.
  PUBMED    34343484
REFERENCE   2  (bases 1 to 9779)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.chembiol.2021.07.011"; journalName: "Cell Chem Biol"; 
            date: "2022-02-17- 17"; volume: "29"; issue: "2"; pages: 
            "321-327.e4"
COMMENT     Sequence Label: pET28-His-MBP-NLS-RfxCas13d-NLS-HA
FEATURES             Location/Qualifiers
     source          1..9779
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     246..265
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     349..368
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        349..367
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    368..392
                     /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)."
     RBS             407..429
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             526..1623
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="KIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLEE
                     KFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKLI
                     AYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIAA
                     DGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGET
                     AMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFLE
                     NYLLTDEGLEAVNKDKPLGAVALKSYEEELVKDPRIAATMENAQKGEIMPNIPQMSAFW
                     YAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     1597..1620
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1960..1980
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1990..4887
                     /codon_start=1
                     /product="RNA-targeting Type VI CRISPR protein from 
                     Ruminococcus flavefaciens strain XPD3002 (Konermann et al.,
                     2018)"
                     /label=RfxCas13d
                     /translation="IEKKKSFAKGMGVKSTLVSGSKVYMTTFAEGSDARLEKIVEGDSI
                     RSVNEGEAFSAEMADKNAGYKIGNAKFSHPKGYAVVANNPLYTGPVQQDMLGLKETLEK
                     RYFGESADGNDNICIQVIHNILDIEKILAEYITNAAYAVNNISGLDKDIIGFGKFSTVY
                     TYDEFKDPEHHRAAFNNNDKLINAIKAQYDEFDNFLDNPRLGYFGQAFFSKEGRNYIIN
                     YGNECYDILALLSGLRHWVVHNNEEESRISRTWLYNLDKNLDNEYISTLNYLYDRITNE
                     LTNSFSKNSAANVNYIAETLGINPAEFAEQYFRFSIMKEQKNLGFNITKLREVMLDRKD
                     MSEIRKNHKVFDSIRTKVYTMMDFVIYRYYIEEDAKVAAANKSLPDNEKSLSEKDIFVI
                     NLRGSFNDDQKDALYYDEANRIWRKLENIMHNIKEFRGNKTREYKKKDAPRLPRILPAG
                     RDVSAFSKLMYALTMFLDGKEINDLLTTLINKFDNIQSFLKVMPLIGVNAKFVEEYAFF
                     KDSAKIADELRLIKSFARMGEPIADARRAMYIDAIRILGTNLSYDELKALADTFSLDEN
                     GNKLKKGKHGMRNFIINNVISNKRFHYLIRYGDPAHLHEIAKNEAVVKFVLGRIADIQK
                     KQGQNGKNQIDRYYETCIGKDKGKSVSEKVDALTKIITGMNYDQFDKKRSVIEDTGREN
                     AEREKFKKIISLYLTVIYHILKNIVNINARYVIGFHCVERDAQLYKEKGYDINLKKLEE
                     KGFSSVTKLCAGIDETAPDKRKDVEKEMAERAKESIDSLESANPKLYANYIKYSDEKKA
                     EEFTRQINREKAKTALNAYLRNTKWNVIIREDLLRIDNKTCTLFRNKAVHLEVARYVHA
                     YINDIAEVNSYFQLYHYIMQRIIMNERYEKSSGKVSEYFDAVNDEKKYNDRLLKLLCVP
                     FGYCIPRFKNLSIEALFDRNEAAKFDKEKKKVSGNS"
     CDS             4897..4917
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4927..4953
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             4992..5009
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(5062..5080)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      5076..5123
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      5160..5615
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5247..5266)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5457..5478
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(5707..6522)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(5707..6522)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     6430..6449
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     6514..6533
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      6644..7232
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7133..7152
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7386..7403
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    7418..7557
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     7643..7665
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(7659..7850)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(8659..9741)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     9703..9722
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(join(9742..9779,1..40))
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc cgggatctcg acgctctccc
       61 ttatgcgact cctgcattag gaagcagccc agtagtaggt tgaggccgtt gagcaccgcc
      121 gccgcaagga atggtgcatg caaggagatg gcgcccaaca gtcccccggc cacggggcct
      181 gccaccatac ccacgccgaa acaagcgctc atgagcccga agtggcgagc ccgatcttcc
      241 ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac ctgtggcgcc ggtgatgccg
      301 gccacgatgc gtccggcgta gaggatcgag atctcgatcc cgcgaaatta atacgactca
      361 ctatagggga attgtgagcg gataacaatt cccctctaga aataattttg tttaacttta
      421 agaaggagat ataccatgaa aatcgaagaa agtaaacatc atcatcacag tggtcatcac
      481 catactggtc accaccatca tagtggcagc caccatcaca ccggcaaaat cgaagaaggt
      541 aaactggtaa tctggattaa cggcgataaa ggctataacg gtctcgctga agtcggtaag
      601 aaattcgaga aagataccgg aattaaagtc accgttgagc atccggataa actggaagag
      661 aaattcccac aggttgcggc aactggcgat ggccctgaca ttatcttctg ggcacacgac
      721 cgctttggtg gctacgctca atctggcctg ttggctgaaa tcaccccgga caaagcgttc
      781 caggacaagc tgtatccgtt tacctgggat gccgtacgtt acaacggcaa gctgattgct
      841 tacccgatcg ctgttgaagc gttatcgctg atttataaca aagatctgct gccgaacccg
      901 ccaaaaacct gggaagagat cccggcgctg gataaagaac tgaaagcgaa aggtaagagc
      961 gcgctgatgt tcaacctgca agaaccgtac ttcacctggc cgctgattgc tgctgacggg
     1021 ggttatgcgt tcaagtatga aaacggcaag tacgacatta aagacgtggg cgtggataac
     1081 gctggcgcga aagcgggtct gaccttcctg gttgacctga ttaaaaacaa acacatgaat
     1141 gcagacaccg attactccat cgcagaagct gcctttaata aaggcgaaac agcgatgacc
     1201 atcaacggcc cgtgggcatg gtccaacatc gacaccagca aagtgaatta tggtgtaacg
     1261 gtactgccga ccttcaaggg tcaaccatcc aaaccgttcg ttggcgtgct gagcgcaggt
     1321 attaacgccg ccagtccgaa caaagagctg gcaaaagagt tcctcgaaaa ctatctgctg
     1381 actgatgaag gtctggaagc ggttaataaa gacaaaccgc tgggtgccgt agcgctgaag
     1441 tcttacgagg aagagttggt gaaagatccg cgtattgccg ccactatgga aaacgcccag
     1501 aaaggtgaaa tcatgccgaa catcccgcag atgtccgctt tctggtatgc cgtgcgtact
     1561 gcggtgatca acgccgccag cggtcgtcag actgtcgatg aagccctgaa agacgcgcag
     1621 actactagtg gcagcaataa tggcagcaat aactcgaata acagctcagc tgcgggtgga
     1681 gaagaggata aaaaaccggc aggcggggag ggaggtggag cacatatcaa cctgaaagtg
     1741 aagggccaag acgggaacga ggtttttttt cgtatcaaac gcagcacaca gctgaagaaa
     1801 ttaatgaacg cctactgtga tcgccagagc gtggacatga cggctattgc tttcttattt
     1861 gatggtcgtc gccttcgtgc ggaacaaacg cctgatgaac ttgagatgga ggatggggat
     1921 gaaattgatg cgatgctgca ccaaaccggt gggatgagcc cgaagaagaa gcgtaaggtt
     1981 gaggcgagca ttgagaagaa gaagagcttt gcgaaaggca tgggcgtgaa gagcaccctg
     2041 gttagcggta gcaaagtgta catgaccacc tttgcggaag gcagcgacgc gcgtctggaa
     2101 aagatcgttg agggtgatag cattcgtagc gtgaacgaag gtgaagcgtt cagcgcggag
     2161 atggcggaca aaaacgcggg ttataagatc ggcaacgcga aatttagcca cccgaagggt
     2221 tatgcggtgg ttgcgaacaa cccgctgtac accggtccgg ttcagcaaga catgctgggc
     2281 ctgaaagaaa ccctggagaa gcgttacttc ggtgaaagcg cggacggcaa cgataacatc
     2341 tgcattcagg tgatccacaa cattctggac atcgaaaaaa ttctggcgga gtatatcacc
     2401 aacgcggcgt acgcggttaa caacattagc ggcctggaca aagacatcat tggtttcggc
     2461 aagtttagca ccgtgtacac ctatgacgaa ttcaaggatc cggagcacca ccgtgcggcg
     2521 tttaacaaca acgacaaact gatcaacgcg attaaggcgc agtacgacga gttcgataac
     2581 tttctggata acccgcgtct gggttatttc ggccaagcgt tctttagcaa agaaggtcgt
     2641 aactatatca ttaactacgg caacgagtgc tacgacattc tggcgctgct gagcggtctg
     2701 cgtcactggg tggttcacaa caacgaggaa gagagccgta tcagccgtac ctggctgtat
     2761 aacctggaca aaaacctgga taacgaatac atcagcaccc tgaactacct gtatgatcgt
     2821 attaccaacg agctgaccaa cagcttcagc aagaacagcg cggcgaacgt taactacatc
     2881 gcggaaaccc tgggtattaa cccggcggaa ttcgcggagc agtatttccg ttttagcatc
     2941 atgaaagagc aaaagaacct gggctttaac attaccaaac tgcgtgaagt tatgctggac
     3001 cgtaaggata tgagcgagat ccgtaaaaac cacaaggtgt tcgatagcat tcgtaccaag
     3061 gtttatacca tgatggactt tgtgatctac cgttactata ttgaagagga tgcgaaagtt
     3121 gcggcggcga acaagagcct gccggacaac gagaagagcc tgagcgagaa ggacatcttc
     3181 gtgattaacc tgcgtggtag ctttaacgac gatcagaaag acgcgctgta ctatgatgaa
     3241 gcgaaccgta tctggcgtaa gctggagaac atcatgcaca acattaaaga attccgtggc
     3301 aacaagaccc gtgagtacaa gaaaaaggac gcgccgcgtc tgccgcgtat cctgccggcg
     3361 ggtcgtgatg ttagcgcgtt cagcaaactg atgtatgcgc tgaccatgtt tctggacggc
     3421 aaggagatta acgatctgct gaccaccctg atcaacaaat tcgacaacat tcaaagcttt
     3481 ctgaaggtga tgccgctgat cggtgttaac gcgaaattcg tggaagagta cgcgttcttt
     3541 aaagacagcg cgaagatcgc ggatgaactg cgtctgatta agagctttgc gcgtatgggt
     3601 gaaccgattg cggatgcgcg tcgtgcgatg tacattgatg cgatccgtat tctgggcacc
     3661 aacctgagct atgacgaact gaaagcgctg gcggacacct ttagcctgga tgagaacggt
     3721 aacaaactga aaaagggtaa acacggcatg cgtaacttca tcattaacaa cgttatcagc
     3781 aacaagcgtt ttcactatct gattcgttac ggtgacccgg cgcacctgca cgaaatcgcg
     3841 aaaaacgagg cggtggttaa gttcgtgctg ggccgtatcg cggatattca gaaaaagcag
     3901 ggtcaaaacg gcaaaaacca aatcgaccgt tactatgaaa cctgcattgg caaggataaa
     3961 ggcaagagcg tgagcgagaa agttgacgcg ctgaccaaga tcattaccgg tatgaactat
     4021 gaccagttcg ataaaaagcg tagcgtgatc gaagacaccg gccgtgagaa cgcggaacgt
     4081 gagaagttca aaaagatcat cagcctgtat ctgaccgtta tctaccacat tctgaagaac
     4141 atcgtgaaca ttaacgcgcg ttacgttatt ggtttccact gcgtggaacg tgatgcgcaa
     4201 ctgtataaag agaagggcta cgacatcaac ctgaaaaagc tggaagagaa aggttttagc
     4261 agcgttacca aactgtgcgc gggtattgat gaaaccgcgc cggacaagcg taaggatgtg
     4321 gaaaaagaga tggcggaacg tgcgaaggag agcatcgaca gcctggagag cgcgaacccg
     4381 aaactgtacg cgaactatat taagtacagc gatgagaaga aggcggaaga gttcacccgt
     4441 cagatcaacc gtgagaaagc gaagaccgcg ctgaacgcgt acctgcgtaa caccaagtgg
     4501 aacgttatca ttcgtgagga cctgctgcgt atcgataaca aaacctgcac cctgttccgt
     4561 aacaaggcgg tgcacctgga agttgcgcgt tatgtgcacg cgtacatcaa cgacattgcg
     4621 gaagtgaaca gctactttca gctgtaccac tatatcatgc aacgtatcat tatgaacgaa
     4681 cgttatgaga aaagcagcgg caaggttagc gaatacttcg acgcggtgaa cgatgagaaa
     4741 aagtataacg atcgtctgct gaaactgctg tgcgttccgt tcggctactg catcccgcgt
     4801 tttaagaacc tgagcattga agcgctgttc gaccgtaacg aggcggcgaa atttgataag
     4861 gagaagaaga aggtgagcgg taacagcggc agcggcccga agaagaagcg taaggttgcg
     4921 gcggcgtacc cgtatgatgt tccggattat gcgtaacgag ctccgtcgac aagcttgcgg
     4981 ccgcactcga gcaccaccac caccaccact gagatccggc tgctaacaaa gcccgaaagg
     5041 aagctgagtt ggctgctgcc accgctgagc aataactagc ataacccctt ggggcctcta
     5101 aacgggtctt gaggggtttt ttgctgaaag gaggaactat atccggattg gcgaatggga
     5161 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     5221 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     5281 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     5341 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     5401 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     5461 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     5521 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     5581 cgcgaatttt aacaaaatat taacgcttac aatttaggtg gcacttttcg gggaaatgtg
     5641 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaat
     5701 taattcttag aaaaactcat cgagcatcaa atgaaactgc aatttattca tatcaggatt
     5761 atcaatacca tatttttgaa aaagccgttt ctgtaatgaa ggagaaaact caccgaggca
     5821 gttccatagg atggcaagat cctggtatcg gtctgcgatt ccgactcgtc caacatcaat
     5881 acaacctatt aatttcccct cgtcaaaaat aaggttatca agtgagaaat caccatgagt
     5941 gacgactgaa tccggtgaga atggcaaaag tttatgcatt tctttccaga cttgttcaac
     6001 aggccagcca ttacgctcgt catcaaaatc actcgcatca accaaaccgt tattcattcg
     6061 tgattgcgcc tgagcgagac gaaatacgcg atcgctgtta aaaggacaat tacaaacagg
     6121 aatcgaatgc aaccggcgca ggaacactgc cagcgcatca acaatatttt cacctgaatc
     6181 aggatattct tctaatacct ggaatgctgt tttcccgggg atcgcagtgg tgagtaacca
     6241 tgcatcatca ggagtacgga taaaatgctt gatggtcgga agaggcataa attccgtcag
     6301 ccagtttagt ctgaccatct catctgtaac atcattggca acgctacctt tgccatgttt
     6361 cagaaacaac tctggcgcat cgggcttccc atacaatcga tagattgtcg cacctgattg
     6421 cccgacatta tcgcgagccc atttataccc atataaatca gcatccatgt tggaatttaa
     6481 tcgcggccta gagcaagacg tttcccgttg aatatggctc ataacacccc ttgtattact
     6541 gtttatgtaa gcagacagtt ttattgttca tgaccaaaat cccttaacgt gagttttcgt
     6601 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     6661 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     6721 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     6781 caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     6841 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     6901 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     6961 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     7021 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     7081 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     7141 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     7201 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     7261 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg
     7321 tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg
     7381 agcgcagcga gtcagtgagc gaggaagcgg aagagcgcct gatgcggtat tttctcctta
     7441 cgcatctgtg cggtatttca caccgcaatg gtgcactctc agtacaatct gctcttatgc
     7501 cgcatagtta agccagtata cactccgcta tcgctacgtg actgggtcat ggctgcgccc
     7561 cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct
     7621 tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca
     7681 ccgaaacgcg cgaggcagct gcggtaaagc tcatcagcgt ggtcgtgaag cgattcacag
     7741 atgtctgcct gttcatccgc gtccagctcg ttgagtttct ccagaagcgt taatgtctgg
     7801 cttctgataa agcgggccat gttaagggcg gttttttcct gtttggtcac tgatgcctcc
     7861 gtgtaagggg gatttctgtt catgggggta atgataccga tgaaacgaga gaggatgctc
     7921 acgatacggg ttactgatga tgaacatgcc cggttactgg aacgttgtga gggtaaacaa
     7981 ctggcggtat ggatgcggcg ggaccagaga aaaatcactc agggtcaatg ccagcgcttc
     8041 gttaatacag atgtaggtgt tccacagggt agccagcagc atcctgcgat gcagatccgg
     8101 aacataatgg tgcagggcgc tgacttccgc gtttccagac tttacgaaac acggaaaccg
     8161 aagaccattc atgttgttgc tcaggtcgca gacgttttgc agcagcagtc gcttcacgtt
     8221 cgctcgcgta tcggtgattc attctgctaa ccagtaaggc aaccccgcca gcctagccgg
     8281 gtcctcaacg acaggagcac gatcatgcgc acccgtgggg ccgccatgcc ggcgataatg
     8341 gcctgcttct cgccgaaacg tttggtggcg ggaccagtga cgaaggcttg agcgagggcg
     8401 tgcaagattc cgaataccgc aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg
     8461 tcctcgccga aaatgaccca gagcgctgcc ggcacctgtc ctacgagttg catgataaag
     8521 aagacagtca taagtgcggc gacgatagtc atgccccgcg cccaccggaa ggagctgact
     8581 gggttgaagg ctctcaaggg catcggtcga gatcccggtg cctaatgagt gagctaactt
     8641 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     8701 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggtggt
     8761 ttttcttttc accagtgaga cgggcaacag ctgattgccc ttcaccgcct ggccctgaga
     8821 gagttgcagc aagcggtcca cgctggtttg ccccagcagg cgaaaatcct gtttgatggt
     8881 ggttaacggc gggatataac atgagctgtc ttcggtatcg tcgtatccca ctaccgagat
     8941 atccgcacca acgcgcagcc cggactcggt aatggcgcgc attgcgccca gcgccatctg
     9001 atcgttggca accagcatcg cagtgggaac gatgccctca ttcagcattt gcatggtttg
     9061 ttgaaaaccg gacatggcac tccagtcgcc ttcccgttcc gctatcggct gaatttgatt
     9121 gcgagtgaga tatttatgcc agccagccag acgcagacgc gccgagacag aacttaatgg
     9181 gcccgctaac agcgcgattt gctggtgacc caatgcgacc agatgctcca cgcccagtcg
     9241 cgtaccgtct tcatgggaga aaataatact gttgatgggt gtctggtcag agacatcaag
     9301 aaataacgcc ggaacattag tgcaggcagc ttccacagca atggcatcct ggtcatccag
     9361 cggatagtta atgatcagcc cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt
     9421 acaggcttcg acgccgcttc gttctaccat cgacaccacc acgctggcac ccagttgatc
     9481 ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca gactggaggt
     9541 ggcaacgcca atcagcaacg actgtttgcc cgccagttgt tgtgccacgc ggttgggaat
     9601 gtaattcagc tccgccatcg ccgcttccac tttttcccgc gttttcgcag aaacgtggct
     9661 ggcctggttc accacgcggg aaacggtctg ataagagaca ccggcatact ctgcgacatc
     9721 gtataacgtt actggtttca cattcaccac cctgaattga ctctcttccg ggcgctatc
//
