LOCUS       sequence-438877-        9286 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  SPNS1-targeting sgRNA in PX458 vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9286)
  AUTHORS   Scharenberg SG, Dong W, Ghoochani A, Nyame K, Levin-Konigsberg R, 
            Krishnan AR, Rawat ES, Spees K, Bassik MC, Abu-Remaileh M
  TITLE     An SPNS1-dependent lysosomal lipid transport pathway that enables 
            cell survival under choline limitation.
  JOURNAL   Sci Adv. 2023 Apr 21;9(16):eadf8966. doi: 10.1126/sciadv.adf8966. 
            Epub 2023 Apr 19.
  PUBMED    37075117
REFERENCE   2  (bases 1 to 9286)
  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"; journalName: "Sci Adv."; 
            date: "2023-04-21"; volume: "9(16):eadf8966. doi"; pages: " 
            10.1126/sciadv.adf8966. Epub 2023 Apr 19"
COMMENT     Sequence Label: PX458-SPNS1-sg1
FEATURES             Location/Qualifiers
     source          1..9286
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..249
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1..21
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     172..191
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        268..343
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        440..725
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        727..1002
                     /label=chicken beta-actin promoter
     intron          1003..1230
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      1242..1251
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1251..1316
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1323..1343
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1368..5468
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             1368..5468
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             5469..5516
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             5532..5585
                     /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             join(5586..5588,5589..6299)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(5586..5588,5589..6299)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(5889..5908)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6236..6257
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(6327..6344)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6333..6540
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   6549..6689
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   6549..6678
                     /label=AAV2 ITR
     rep_origin      6764..7219
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6851..6870)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7061..7082
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(7236..7255)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7355..7377
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7415..7433)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7501..7605
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7606..7674,7675..8466)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(7824..7843)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8637..9225
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     9126..9145
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
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 gtcggccaca aagaggtgtt ttagagctag aaatagcaag ttaaaataag
      301 gctagtccgt tatcaacttg aaaaagtggc accgagtcgg tgcttttttg ttttagagct
      361 agaaatagca agttaaaata aggctagtcc gtttttagcg cgtgcgccaa ttctgcagac
      421 aaatggctct agaggtaccc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      481 ccaacgaccc ccgcccattg acgtcaatag taacgccaat agggactttc cattgacgtc
      541 aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc
      601 caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tgtgcccagt
      661 acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta
      721 ccatggtcga ggtgagcccc acgttctgct tcactctccc catctccccc ccctccccac
      781 ccccaatttt gtatttattt attttttaat tattttgtgc agcgatgggg gcgggggggg
      841 ggggggcgcg cgccaggcgg ggcggggcgg ggcgaggggc ggggcggggc gaggcggaga
      901 ggtgcggcgg cagccaatca gagcggcgcg ctccgaaagt ttccttttat ggcgaggcgg
      961 cggcggcggc ggccctataa aaagcgaagc gcgcggcggg cgggagtcgc tgcgcgctgc
     1021 cttcgccccg tgccccgctc cgccgccgcc tcgcgccgcc cgccccggct ctgactgacc
     1081 gcgttactcc cacaggtgag cgggcgggac ggcccttctc ctccgggctg taattagctg
     1141 agcaagaggt aagggtttaa gggatggttg gttggtgggg tattaatgtt taattacctg
     1201 gagcacctgc ctgaaatcac tttttttcag gttggaccgg tgccaccatg gactataagg
     1261 accacgacgg agactacaag gatcatgata ttgattacaa agacgatgac gataagatgg
     1321 ccccaaagaa gaagcggaag gtcggtatcc acggagtccc agcagccgac aagaagtaca
     1381 gcatcggcct ggacatcggc accaactctg tgggctgggc cgtgatcacc gacgagtaca
     1441 aggtgcccag caagaaattc aaggtgctgg gcaacaccga ccggcacagc atcaagaaga
     1501 acctgatcgg agccctgctg ttcgacagcg gcgaaacagc cgaggccacc cggctgaaga
     1561 gaaccgccag aagaagatac accagacgga agaaccggat ctgctatctg caagagatct
     1621 tcagcaacga gatggccaag gtggacgaca gcttcttcca cagactggaa gagtccttcc
     1681 tggtggaaga ggataagaag cacgagcggc accccatctt cggcaacatc gtggacgagg
     1741 tggcctacca cgagaagtac cccaccatct accacctgag aaagaaactg gtggacagca
     1801 ccgacaaggc cgacctgcgg ctgatctatc tggccctggc ccacatgatc aagttccggg
     1861 gccacttcct gatcgagggc gacctgaacc ccgacaacag cgacgtggac aagctgttca
     1921 tccagctggt gcagacctac aaccagctgt tcgaggaaaa ccccatcaac gccagcggcg
     1981 tggacgccaa ggccatcctg tctgccagac tgagcaagag cagacggctg gaaaatctga
     2041 tcgcccagct gcccggcgag aagaagaatg gcctgttcgg aaacctgatt gccctgagcc
     2101 tgggcctgac ccccaacttc aagagcaact tcgacctggc cgaggatgcc aaactgcagc
     2161 tgagcaagga cacctacgac gacgacctgg acaacctgct ggcccagatc ggcgaccagt
     2221 acgccgacct gtttctggcc gccaagaacc tgtccgacgc catcctgctg agcgacatcc
     2281 tgagagtgaa caccgagatc accaaggccc ccctgagcgc ctctatgatc aagagatacg
     2341 acgagcacca ccaggacctg accctgctga aagctctcgt gcggcagcag ctgcctgaga
     2401 agtacaaaga gattttcttc gaccagagca agaacggcta cgccggctac attgacggcg
     2461 gagccagcca ggaagagttc tacaagttca tcaagcccat cctggaaaag atggacggca
     2521 ccgaggaact gctcgtgaag ctgaacagag aggacctgct gcggaagcag cggaccttcg
     2581 acaacggcag catcccccac cagatccacc tgggagagct gcacgccatt ctgcggcggc
     2641 aggaagattt ttacccattc ctgaaggaca accgggaaaa gatcgagaag atcctgacct
     2701 tccgcatccc ctactacgtg ggccctctgg ccaggggaaa cagcagattc gcctggatga
     2761 ccagaaagag cgaggaaacc atcaccccct ggaacttcga ggaagtggtg gacaagggcg
     2821 cttccgccca gagcttcatc gagcggatga ccaacttcga taagaacctg cccaacgaga
     2881 aggtgctgcc caagcacagc ctgctgtacg agtacttcac cgtgtataac gagctgacca
     2941 aagtgaaata cgtgaccgag ggaatgagaa agcccgcctt cctgagcggc gagcagaaaa
     3001 aggccatcgt ggacctgctg ttcaagacca accggaaagt gaccgtgaag cagctgaaag
     3061 aggactactt caagaaaatc gagtgcttcg actccgtgga aatctccggc gtggaagatc
     3121 ggttcaacgc ctccctgggc acataccacg atctgctgaa aattatcaag gacaaggact
     3181 tcctggacaa tgaggaaaac gaggacattc tggaagatat cgtgctgacc ctgacactgt
     3241 ttgaggacag agagatgatc gaggaacggc tgaaaaccta tgcccacctg ttcgacgaca
     3301 aagtgatgaa gcagctgaag cggcggagat acaccggctg gggcaggctg agccggaagc
     3361 tgatcaacgg catccgggac aagcagtccg gcaagacaat cctggatttc ctgaagtccg
     3421 acggcttcgc caacagaaac ttcatgcagc tgatccacga cgacagcctg acctttaaag
     3481 aggacatcca gaaagcccag gtgtccggcc agggcgatag cctgcacgag cacattgcca
     3541 atctggccgg cagccccgcc attaagaagg gcatcctgca gacagtgaag gtggtggacg
     3601 agctcgtgaa agtgatgggc cggcacaagc ccgagaacat cgtgatcgaa atggccagag
     3661 agaaccagac cacccagaag ggacagaaga acagccgcga gagaatgaag cggatcgaag
     3721 agggcatcaa agagctgggc agccagatcc tgaaagaaca ccccgtggaa aacacccagc
     3781 tgcagaacga gaagctgtac ctgtactacc tgcagaatgg gcgggatatg tacgtggacc
     3841 aggaactgga catcaaccgg ctgtccgact acgatgtgga ccatatcgtg cctcagagct
     3901 ttctgaagga cgactccatc gacaacaagg tgctgaccag aagcgacaag aaccggggca
     3961 agagcgacaa cgtgccctcc gaagaggtcg tgaagaagat gaagaactac tggcggcagc
     4021 tgctgaacgc caagctgatt acccagagaa agttcgacaa tctgaccaag gccgagagag
     4081 gcggcctgag cgaactggat aaggccggct tcatcaagag acagctggtg gaaacccggc
     4141 agatcacaaa gcacgtggca cagatcctgg actcccggat gaacactaag tacgacgaga
     4201 atgacaagct gatccgggaa gtgaaagtga tcaccctgaa gtccaagctg gtgtccgatt
     4261 tccggaagga tttccagttt tacaaagtgc gcgagatcaa caactaccac cacgcccacg
     4321 acgcctacct gaacgccgtc gtgggaaccg ccctgatcaa aaagtaccct aagctggaaa
     4381 gcgagttcgt gtacggcgac tacaaggtgt acgacgtgcg gaagatgatc gccaagagcg
     4441 agcaggaaat cggcaaggct accgccaagt acttcttcta cagcaacatc atgaactttt
     4501 tcaagaccga gattaccctg gccaacggcg agatccggaa gcggcctctg atcgagacaa
     4561 acggcgaaac cggggagatc gtgtgggata agggccggga ttttgccacc gtgcggaaag
     4621 tgctgagcat gccccaagtg aatatcgtga aaaagaccga ggtgcagaca ggcggcttca
     4681 gcaaagagtc tatcctgccc aagaggaaca gcgataagct gatcgccaga aagaaggact
     4741 gggaccctaa gaagtacggc ggcttcgaca gccccaccgt ggcctattct gtgctggtgg
     4801 tggccaaagt ggaaaagggc aagtccaaga aactgaagag tgtgaaagag ctgctgggga
     4861 tcaccatcat ggaaagaagc agcttcgaga agaatcccat cgactttctg gaagccaagg
     4921 gctacaaaga agtgaaaaag gacctgatca tcaagctgcc taagtactcc ctgttcgagc
     4981 tggaaaacgg ccggaagaga atgctggcct ctgccggcga actgcagaag ggaaacgaac
     5041 tggccctgcc ctccaaatat gtgaacttcc tgtacctggc cagccactat gagaagctga
     5101 agggctcccc cgaggataat gagcagaaac agctgtttgt ggaacagcac aagcactacc
     5161 tggacgagat catcgagcag atcagcgagt tctccaagag agtgatcctg gccgacgcta
     5221 atctggacaa agtgctgtcc gcctacaaca agcaccggga taagcccatc agagagcagg
     5281 ccgagaatat catccacctg tttaccctga ccaatctggg agcccctgcc gccttcaagt
     5341 actttgacac caccatcgac cggaagaggt acaccagcac caaagaggtg ctggacgcca
     5401 ccctgatcca ccagagcatc accggcctgt acgagacacg gatcgacctg tctcagctgg
     5461 gaggcgacaa aaggccggcg gccacgaaaa aggccggcca ggcaaaaaag aaaaaggaat
     5521 tcggcagtgg agagggcaga ggaagtctgc taacatgcgg tgacgtcgag gagaatcctg
     5581 gcccagtgag caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg
     5641 acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct
     5701 acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca
     5761 ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc gaccacatga
     5821 agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct
     5881 tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc
     5941 tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc
     6001 acaagctgga gtacaactac aacagccaca acgtctatat catggccgac aagcagaaga
     6061 acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc gtgcagctcg
     6121 ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc
     6181 actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc gatcacatgg
     6241 tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagg
     6301 aattctaact agagctcgct gatcagcctc gactgtgcct tctagttgcc agccatctgt
     6361 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     6421 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
     6481 tggggtgggg caggacagca agggggagga ttgggaagag aatagcaggc atgctgggga
     6541 gcggccgcag gaacccctag tgatggagtt ggccactccc tctctgcgcg ctcgctcgct
     6601 cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg cggcctcagt
     6661 gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc tccttacgca
     6721 tctgtgcggt atttcacacc gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc
     6781 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     6841 ttagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     6901 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     6961 gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     7021 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     7081 ggaacaacac tcaactctat ctcgggctat tcttttgatt tataagggat tttgccgatt
     7141 tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     7201 atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga tgccgcatag
     7261 ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     7321 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     7381 tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag
     7441 gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg
     7501 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     7561 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
     7621 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     7681 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
     7741 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
     7801 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg
     7861 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
     7921 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     7981 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     8041 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     8101 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
     8161 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     8221 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     8281 ggctggttta ttgctgataa atctggagcc ggtgagcgtg gaagccgcgg tatcattgca
     8341 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     8401 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     8461 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     8521 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     8581 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     8641 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     8701 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     8761 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     8821 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     8881 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     8941 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     9001 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     9061 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     9121 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     9181 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     9241 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgt
//
