LOCUS       sequence-322788-       11589 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  lentiviral vector expressing Cas9 and an sgRNA targeting GLS.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11589)
  AUTHORS   Rossiter NJ, Huggler KS, Adelmann CH, Keys HR, Soens RW, Sabatini 
            DM, Cantor JR
  TITLE     CRISPR screens in physiologic medium reveal conditionally essential 
            genes in human cells.
  JOURNAL   Cell Metab. 2021 Feb 23. pii: S1550-4131(21)00061-9. doi: 
            10.1016/j.cmet.2021.02.005.
  PUBMED    33651980
REFERENCE   2  (bases 1 to 11589)
  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: "Cell Metab.
            2021 Feb 23. pii: S1550-4131(21)00061-9. doi: 
            10.1016/j.cmet.2021.02.005."
COMMENT     Sequence Label: pLentiCRISPR-v1-sgGLS_6
FEATURES             Location/Qualifiers
     source          1..11589
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..181
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    228..353
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    846..1079
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1264..1308
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     promoter        1486..1734
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1486..1506
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     1657..1676
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        1755..1830
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_feature    1878..1995
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2064..2275
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      2295..2304
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2304..2327
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2334..2354
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2379..6479
                     /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             2379..6479
                     /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             6480..6527
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             6543..6599
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             6600..7199
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(6600..6619)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7096..7116
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    7215..7803
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7268..7288)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(7686..7697)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             7875..8108
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    8186..8307
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8223..8242)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8277..8296
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      8347..8482
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(8502..8521)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(8503..8521)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(8531..8548)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(8531..8547)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8540..8562)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      8689..9144
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8776..8795)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     8986..9007
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        9170..9274
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(9275..9343,9344..10135)
                     /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(9493..9512)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      10306..10894
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     10795..10814
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     11048..11065
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    11182..11203
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(11218..11223,11224..11241,11242..11248)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11256..11272
                     /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)."
     primer_bind     11261..11283
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     11280..11296
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     11280..11296
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     11315..11335
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        11317..11335
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        11363..11589
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
ORIGIN
        1 gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact agggaaccca
       61 ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg
      121 tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa aatctctagc
      181 agtggcgccc gaacagggac ttgaaagcga aagggaaacc agaggagctc tctcgacgca
      241 ggactcggct tgctgaagcg cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc
      301 caaaaatttt gactagcgga ggctagaagg agagagatgg gtgcgagagc gtcagtatta
      361 agcgggggag aattagatcg cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa
      421 aatataaatt aaaacatata gtatgggcaa gcagggagct agaacgattc gcagttaatc
      481 ctggcctgtt agaaacatca gaaggctgta gacaaatact gggacagcta caaccatccc
      541 ttcagacagg atcagaagaa cttagatcat tatataatac agtagcaacc ctctattgtg
      601 tgcatcaaag gatagagata aaagacacca aggaagcttt agacaagata gaggaagagc
      661 aaaacaaaag taagaccacc gcacagcaag cggccgctga tcttcagacc tggaggagga
      721 gatatgaggg acaattggag aagtgaatta tataaatata aagtagtaaa aattgaacca
      781 ttaggagtag cacccaccaa ggcaaagaga agagtggtgc agagagaaaa aagagcagtg
      841 ggaataggag ctttgttcct tgggttcttg ggagcagcag gaagcactat gggcgcagcg
      901 tcaatgacgc tgacggtaca ggccagacaa ttattgtctg gtatagtgca gcagcagaac
      961 aatttgctga gggctattga ggcgcaacag catctgttgc aactcacagt ctggggcatc
     1021 aagcagctcc aggcaagaat cctggctgtg gaaagatacc taaaggatca acagctcctg
     1081 gggatttggg gttgctctgg aaaactcatt tgcaccactg ctgtgccttg gaatgctagt
     1141 tggagtaata aatctctgga acagatttgg aatcacacga cctggatgga gtgggacaga
     1201 gaaattaaca attacacaag cttaatacac tccttaattg aagaatcgca aaaccagcaa
     1261 gaaaagaatg aacaagaatt attggaatta gataaatggg caagtttgtg gaattggttt
     1321 aacataacaa attggctgtg gtatataaaa ttattcataa tgatagtagg aggcttggta
     1381 ggtttaagaa tagtttttgc tgtactttct atagtgaata gagttaggca gggatattca
     1441 ccattatcgt ttcagaccca cctcccaacc ccgaggggac ccagagaggg cctatttccc
     1501 atgattcctt catatttgca tatacgatac aaggctgtta gagagataat tagaattaat
     1561 ttgactgtaa acacaaagat attagtacaa aatacgtgac gtagaaagta ataatttctt
     1621 gggtagtttg cagttttaaa attatgtttt aaaatggact atcatatgct taccgtaact
     1681 tgaaagtatt tcgatttctt ggctttatat atcttgtgga aaggacgaaa caccgctgcg
     1741 acactggcta atgggtttta gagctagaaa tagcaagtta aaataaggct agtccgttat
     1801 caacttgaaa aagtggcacc gagtcggtgc ttttttgaat tctagatctt gagacaaatg
     1861 gcagtattca tccacaattt taaaagaaaa ggggggattg gggggtacag tgcaggggaa
     1921 agaatagtag acataatagc aacagacata caaactaaag aattacaaaa acaaattaca
     1981 aaaattcaaa attttcgggt ttattacagg gacagcagag atccactttg gcgccggctc
     2041 gagtggctcc ggtgcccgtc agtgggcaga gcgcacatcg cccacagtcc ccgagaagtt
     2101 ggggggaggg gtcggcaatt gaaccggtgc ctagagaagg tggcgcgggg taaactggga
     2161 aagtgatgtc gtgtactggc tccgcctttt tcccgagggt gggggagaac cgtatataag
     2221 tgcagtagtc gccgtgaacg ttctttttcg caacgggttt gccgccagaa cacaggtgtc
     2281 gtgacgcggg atccgccacc atggattaca aagacgatga cgataagatg gccccaaaga
     2341 agaagcggaa ggtcggtatc cacggagtcc cagcagccga caagaagtac agcatcggcc
     2401 tggacatcgg caccaactct gtgggctggg ccgtgatcac cgacgagtac aaggtgccca
     2461 gcaagaaatt caaggtgctg ggcaacaccg accggcacag catcaagaag aacctgatcg
     2521 gagccctgct gttcgacagc ggcgaaacag ccgaggccac ccggctgaag agaaccgcca
     2581 gaagaagata caccagacgg aagaaccgga tctgctatct gcaagagatc ttcagcaacg
     2641 agatggccaa ggtggacgac agcttcttcc acagactgga agagtccttc ctggtggaag
     2701 aggataagaa gcacgagcgg caccccatct tcggcaacat cgtggacgag gtggcctacc
     2761 acgagaagta ccccaccatc taccacctga gaaagaaact ggtggacagc accgacaagg
     2821 ccgacctgcg gctgatctat ctggccctgg cccacatgat caagttccgg ggccacttcc
     2881 tgatcgaggg cgacctgaac cccgacaaca gcgacgtgga caagctgttc atccagctgg
     2941 tgcagaccta caaccagctg ttcgaggaaa accccatcaa cgccagcggc gtggacgcca
     3001 aggccatcct gtctgccaga ctgagcaaga gcagacggct ggaaaatctg atcgcccagc
     3061 tgcccggcga gaagaagaat ggcctgttcg gaaacctgat tgccctgagc ctgggcctga
     3121 cccccaactt caagagcaac ttcgacctgg ccgaggatgc caaactgcag ctgagcaagg
     3181 acacctacga cgacgacctg gacaacctgc tggcccagat cggcgaccag tacgccgacc
     3241 tgtttctggc cgccaagaac ctgtccgacg ccatcctgct gagcgacatc ctgagagtga
     3301 acaccgagat caccaaggcc cccctgagcg cctctatgat caagagatac gacgagcacc
     3361 accaggacct gaccctgctg aaagctctcg tgcggcagca gctgcctgag aagtacaaag
     3421 agattttctt cgaccagagc aagaacggct acgccggcta cattgacggc ggagccagcc
     3481 aggaagagtt ctacaagttc atcaagccca tcctggaaaa gatggacggc accgaggaac
     3541 tgctcgtgaa gctgaacaga gaggacctgc tgcggaagca gcggaccttc gacaacggca
     3601 gcatccccca ccagatccac ctgggagagc tgcacgccat tctgcggcgg caggaagatt
     3661 tttacccatt cctgaaggac aaccgggaaa agatcgagaa gatcctgacc ttccgcatcc
     3721 cctactacgt gggccctctg gccaggggaa acagcagatt cgcctggatg accagaaaga
     3781 gcgaggaaac catcaccccc tggaacttcg aggaagtggt ggacaagggc gcttccgccc
     3841 agagcttcat cgagcggatg accaacttcg ataagaacct gcccaacgag aaggtgctgc
     3901 ccaagcacag cctgctgtac gagtacttca ccgtgtataa cgagctgacc aaagtgaaat
     3961 acgtgaccga gggaatgaga aagcccgcct tcctgagcgg cgagcagaaa aaggccatcg
     4021 tggacctgct gttcaagacc aaccggaaag tgaccgtgaa gcagctgaaa gaggactact
     4081 tcaagaaaat cgagtgcttc gactccgtgg aaatctccgg cgtggaagat cggttcaacg
     4141 cctccctggg cacataccac gatctgctga aaattatcaa ggacaaggac ttcctggaca
     4201 atgaggaaaa cgaggacatt ctggaagata tcgtgctgac cctgacactg tttgaggaca
     4261 gagagatgat cgaggaacgg ctgaaaacct atgcccacct gttcgacgac aaagtgatga
     4321 agcagctgaa gcggcggaga tacaccggct ggggcaggct gagccggaag ctgatcaacg
     4381 gcatccggga caagcagtcc ggcaagacaa tcctggattt cctgaagtcc gacggcttcg
     4441 ccaacagaaa cttcatgcag ctgatccacg acgacagcct gacctttaaa gaggacatcc
     4501 agaaagccca ggtgtccggc cagggcgata gcctgcacga gcacattgcc aatctggccg
     4561 gcagccccgc cattaagaag ggcatcctgc agacagtgaa ggtggtggac gagctcgtga
     4621 aagtgatggg ccggcacaag cccgagaaca tcgtgatcga aatggccaga gagaaccaga
     4681 ccacccagaa gggacagaag aacagccgcg agagaatgaa gcggatcgaa gagggcatca
     4741 aagagctggg cagccagatc ctgaaagaac accccgtgga aaacacccag ctgcagaacg
     4801 agaagctgta cctgtactac ctgcagaatg ggcgggatat gtacgtggac caggaactgg
     4861 acatcaaccg gctgtccgac tacgatgtgg accatatcgt gcctcagagc tttctgaagg
     4921 acgactccat cgacaacaag gtgctgacca gaagcgacaa gaaccggggc aagagcgaca
     4981 acgtgccctc cgaagaggtc gtgaagaaga tgaagaacta ctggcggcag ctgctgaacg
     5041 ccaagctgat tacccagaga aagttcgaca atctgaccaa ggccgagaga ggcggcctga
     5101 gcgaactgga taaggccggc ttcatcaaga gacagctggt ggaaacccgg cagatcacaa
     5161 agcacgtggc acagatcctg gactcccgga tgaacactaa gtacgacgag aatgacaagc
     5221 tgatccggga agtgaaagtg atcaccctga agtccaagct ggtgtccgat ttccggaagg
     5281 atttccagtt ttacaaagtg cgcgagatca acaactacca ccacgcccac gacgcctacc
     5341 tgaacgccgt cgtgggaacc gccctgatca aaaagtaccc taagctggaa agcgagttcg
     5401 tgtacggcga ctacaaggtg tacgacgtgc ggaagatgat cgccaagagc gagcaggaaa
     5461 tcggcaaggc taccgccaag tacttcttct acagcaacat catgaacttt ttcaagaccg
     5521 agattaccct ggccaacggc gagatccgga agcggcctct gatcgagaca aacggcgaaa
     5581 ccggggagat cgtgtgggat aagggccggg attttgccac cgtgcggaaa gtgctgagca
     5641 tgccccaagt gaatatcgtg aaaaagaccg aggtgcagac aggcggcttc agcaaagagt
     5701 ctatcctgcc caagaggaac agcgataagc tgatcgccag aaagaaggac tgggacccta
     5761 agaagtacgg cggcttcgac agccccaccg tggcctattc tgtgctggtg gtggccaaag
     5821 tggaaaaggg caagtccaag aaactgaaga gtgtgaaaga gctgctgggg atcaccatca
     5881 tggaaagaag cagcttcgag aagaatccca tcgactttct ggaagccaag ggctacaaag
     5941 aagtgaaaaa ggacctgatc atcaagctgc ctaagtactc cctgttcgag ctggaaaacg
     6001 gccggaagag aatgctggcc tctgccggcg aactgcagaa gggaaacgaa ctggccctgc
     6061 cctccaaata tgtgaacttc ctgtacctgg ccagccacta tgagaagctg aagggctccc
     6121 ccgaggataa tgagcagaaa cagctgtttg tggaacagca caagcactac ctggacgaga
     6181 tcatcgagca gatcagcgag ttctccaaga gagtgatcct ggccgacgct aatctggaca
     6241 aagtgctgtc cgcctacaac aagcaccggg ataagcccat cagagagcag gccgagaata
     6301 tcatccacct gtttaccctg accaatctgg gagcccctgc cgccttcaag tactttgaca
     6361 ccaccatcga ccggaagagg tacaccagca ccaaagaggt gctggacgcc accctgatcc
     6421 accagagcat caccggcctg tacgagacac ggatcgacct gtctcagctg ggaggcgaca
     6481 agcgtcctgc tgctactaag aaagctggtc aagctaagaa aaagaaagct agcggcagcg
     6541 gcgccaccaa cttcagcctg ctgaagcagg ccggcgacgt ggaggagaac cccggcccca
     6601 tgaccgagta caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac
     6661 gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc gatccggacc
     6721 gccacatcga gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc gggctcgaca
     6781 tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc acgccggaga
     6841 gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg catggccgag ttgagcggtt
     6901 cccggctggc cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg cccaaggagc
     6961 ccgcgtggtt cctggccacc gtcggagtct cgcccgacca ccagggcaag ggtctgggca
     7021 gcgccgtcgt gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg
     7081 agacctccgc gccccgcaac ctccccttct acgagcggct cggcttcacc gtcaccgccg
     7141 acgtcgaggt gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctgaa
     7201 cgcgttaagt cgacaatcaa cctctggatt acaaaatttg tgaaagattg actggtattc
     7261 ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg
     7321 ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg ttgctgtctc
     7381 tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg
     7441 acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc gggactttcg
     7501 ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc cgctgctgga
     7561 caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct
     7621 ttccttggct gctcgcctgt gttgccacct ggattctgcg cgggacgtcc ttctgctacg
     7681 tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg gctctgcggc
     7741 ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat ctccctttgg gccgcctccc
     7801 cgcgtcgact ttaagaccaa tgacttacaa ggcagctgta gatcttagcc actttttaaa
     7861 agaaaagggg ggactggaag ggctaattca ctcccaacga agacaagatc tgctttttgc
     7921 ttgtactggg tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg
     7981 gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg
     8041 tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat
     8101 ctctagcagt acgtatagta gttcatgtca tcttattatt cagtatttat aacttgcaaa
     8161 gaaatgaata tcagagagtg agaggaactt gtttattgca gcttataatg gttacaaata
     8221 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     8281 tttgtccaaa ctcatcaatg tatcttatca tgtctggctc tagctatccc gcccctaact
     8341 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     8401 atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag
     8461 tgaggaggct tttttggagg cctagggacg tacccaattc gccctatagt gagtcgtatt
     8521 acgcgcgctc actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc
     8581 aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc
     8641 gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatgggac gcgccctgta
     8701 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     8761 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     8821 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     8881 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     8941 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     9001 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     9061 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     9121 acaaaatatt aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     9181 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     9241 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     9301 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     9361 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     9421 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac
     9481 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact
     9541 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     9601 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     9661 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     9721 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     9781 agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg
     9841 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     9901 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     9961 tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc
    10021 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
    10081 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc
    10141 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
    10201 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
    10261 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
    10321 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
    10381 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
    10441 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
    10501 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
    10561 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
    10621 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
    10681 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
    10741 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
    10801 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
    10861 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
    10921 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
    10981 tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac
    11041 cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct
    11101 ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc
    11161 gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt
    11221 acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac
    11281 aggaaacagc tatgaccatg attacgccaa gcgcgcaatt aaccctcact aaagggaaca
    11341 aaagctggag ctgcaagctt aatgtagtct tatgcaatac tcttgtagtc ttgcaacatg
    11401 gtaacgatga gttagcaaca tgccttacaa ggagagaaaa agcaccgtgc atgccgattg
    11461 gtggaagtaa ggtggtacga tcgtgcctta ttaggaaggc aacagacggg tctgacatgg
    11521 attggacgaa ccactgaatt gccgcattgc agagatattg tatttaagtg cctagctcga
    11581 tacataaac
//
