LOCUS       sequence-364862-       13013 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  knock out DHODH in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13013)
  AUTHORS   Mao C, Liu X, Zhang Y, Lei G, Yan Y, Lee H, Koppula P, Wu S, Zhuang 
            L, Fang B, Poyurovsky MV, Olszewski K, Gan B
  TITLE     DHODH-mediated ferroptosis defence is a targetable vulnerability in 
            cancer.
  JOURNAL   Nature. 2021 May;593(7860):586-590. doi: 10.1038/s41586-021-03539-7.
            Epub 2021 May 12.
  PUBMED    33981038
REFERENCE   2  (bases 1 to 13013)
  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: 
            "10.1038/s41586-021-03539-7"; journalName: "Nature"; date: 
            "2021-05"; volume: "593"; issue: "7860"; pages: "586-590"
COMMENT     Sequence Label: lentiCRISPR v2-sgDHODH-4
FEATURES             Location/Qualifiers
     source          1..13013
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(20..37)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(191..779)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(271..290)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(950..1741,1742..1810))
                     /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     1573..1592
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1811..1915)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1990..2009)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2181..2560
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2562..2760
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2715..2735
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             2778..2958
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    3005..3130
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    3623..3856
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             4041..4085
                     /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"
     misc_feature    4383..4500
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        4551..4799
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     4551..4571
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     4722..4741
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        4821..4896
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        4958..5169
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      5188..5197
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             5194..9297
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             5194..9297
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             9298..9345
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             9346..9369
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             9379..9435
                     /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             9436..10032
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
                     TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
                     QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
                     PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     9929..9949
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    10048..10636
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(10101..10121)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(10519..10530)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             10708..10941
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(10967..10984)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    10973..11197
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      11243..11671
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(11330..11349)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     11540..11561
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(11680..11700)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        11685..12014
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      11865..12000
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     11927..11946
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        12062..12109
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             12128..12502
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    12632..12753
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(12669..12688)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     12723..12742
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(12802..12818)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(12802..12818)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(12815..12837)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    12826..12842
                     /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(join(12850..12856,12857..12874,12875..12880))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    12895..12916
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
       61 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
      121 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
      181 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
      241 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
      301 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
      361 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
      421 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
      481 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
      541 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
      601 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
      661 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
      721 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
      781 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
      841 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
      901 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
      961 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     1021 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     1081 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     1141 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     1201 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     1261 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     1321 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     1381 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     1441 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     1501 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     1561 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     1621 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     1681 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     1741 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     1801 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     1861 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     1921 ttccccgaaa agtgccacct gacgtcgacg gatcgggaga tctcccgatc ccctatggtg
     1981 cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatctgc tccctgcttg
     2041 tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt
     2101 gaccgacaat tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc ttcgcgatgt
     2161 acgggccaga tatacgcgtt gacattgatt attgactagt tattaatagt aatcaattac
     2221 ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg
     2281 cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc
     2341 catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac
     2401 tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa
     2461 tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac
     2521 ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta
     2581 catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga
     2641 cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa
     2701 ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag
     2761 cgcgttttgc ctgtactggg tctctctggt tagaccagat ctgagcctgg gagctctctg
     2821 gctaactagg gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag
     2881 tgtgtgcccg tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag
     2941 tgtggaaaat ctctagcagt ggcgcccgaa cagggacttg aaagcgaaag ggaaaccaga
     3001 ggagctctct cgacgcagga ctcggcttgc tgaagcgcgc acggcaagag gcgaggggcg
     3061 gcgactggtg agtacgccaa aaattttgac tagcggaggc tagaaggaga gagatgggtg
     3121 cgagagcgtc agtattaagc gggggagaat tagatcgcga tgggaaaaaa ttcggttaag
     3181 gccaggggga aagaaaaaat ataaattaaa acatatagta tgggcaagca gggagctaga
     3241 acgattcgca gttaatcctg gcctgttaga aacatcagaa ggctgtagac aaatactggg
     3301 acagctacaa ccatcccttc agacaggatc agaagaactt agatcattat ataatacagt
     3361 agcaaccctc tattgtgtgc atcaaaggat agagataaaa gacaccaagg aagctttaga
     3421 caagatagag gaagagcaaa acaaaagtaa gaccaccgca cagcaagcgg ccgctgatct
     3481 tcagacctgg aggaggagat atgagggaca attggagaag tgaattatat aaatataaag
     3541 tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc aaagagaaga gtggtgcaga
     3601 gagaaaaaag agcagtggga ataggagctt tgttccttgg gttcttggga gcagcaggaa
     3661 gcactatggg cgcagcgtca atgacgctga cggtacaggc cagacaatta ttgtctggta
     3721 tagtgcagca gcagaacaat ttgctgaggg ctattgaggc gcaacagcat ctgttgcaac
     3781 tcacagtctg gggcatcaag cagctccagg caagaatcct ggctgtggaa agatacctaa
     3841 aggatcaaca gctcctgggg atttggggtt gctctggaaa actcatttgc accactgctg
     3901 tgccttggaa tgctagttgg agtaataaat ctctggaaca gatttggaat cacacgacct
     3961 ggatggagtg ggacagagaa attaacaatt acacaagctt aatacactcc ttaattgaag
     4021 aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt ggaattagat aaatgggcaa
     4081 gtttgtggaa ttggtttaac ataacaaatt ggctgtggta tataaaatta ttcataatga
     4141 tagtaggagg cttggtaggt ttaagaatag tttttgctgt actttctata gtgaatagag
     4201 ttaggcaggg atattcacca ttatcgtttc agacccacct cccaaccccg aggggacccg
     4261 acaggcccga aggaatagaa gaagaaggtg gagagagaga cagagacaga tccattcgat
     4321 tagtgaacgg atcggcactg cgtgcgccaa ttctgcagac aaatggcagt attcatccac
     4381 aattttaaaa gaaaaggggg gattgggggg tacagtgcag gggaaagaat agtagacata
     4441 atagcaacag acatacaaac taaagaatta caaaaacaaa ttacaaaaat tcaaaatttt
     4501 cgggtttatt acagggacag cagagatcca gtttggttaa ttaaggtacc gagggcctat
     4561 ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag ataattagaa
     4621 ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga aagtaataat
     4681 ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat atgcttaccg
     4741 taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga cgaaacaccg
     4801 tgagttgata aatcccggag gttttagagc tagaaatagc aagttaaaat aaggctagtc
     4861 cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt ttgaattcgc tagctaggtc
     4921 ttgaaaggag tgggaattgg ctccggtgcc cgtcagtggg cagagcgcac atcgcccaca
     4981 gtccccgaga agttgggggg aggggtcggc aattgatccg gtgcctagag aaggtggcgc
     5041 ggggtaaact gggaaagtga tgtcgtgtac tggctccgcc tttttcccga gggtggggga
     5101 gaaccgtata taagtgcagt agtcgccgtg aacgttcttt ttcgcaacgg gtttgccgcc
     5161 agaacacagg accggttcta gagcgctgcc accatggaca agaagtacag catcggcctg
     5221 gacatcggca ccaactctgt gggctgggcc gtgatcaccg acgagtacaa ggtgcccagc
     5281 aagaaattca aggtgctggg caacaccgac cggcacagca tcaagaagaa cctgatcgga
     5341 gccctgctgt tcgacagcgg cgaaacagcc gaggccaccc ggctgaagag aaccgccaga
     5401 agaagataca ccagacggaa gaaccggatc tgctatctgc aagagatctt cagcaacgag
     5461 atggccaagg tggacgacag cttcttccac agactggaag agtccttcct ggtggaagag
     5521 gataagaagc acgagcggca ccccatcttc ggcaacatcg tggacgaggt ggcctaccac
     5581 gagaagtacc ccaccatcta ccacctgaga aagaaactgg tggacagcac cgacaaggcc
     5641 gacctgcggc tgatctatct ggccctggcc cacatgatca agttccgggg ccacttcctg
     5701 atcgagggcg acctgaaccc cgacaacagc gacgtggaca agctgttcat ccagctggtg
     5761 cagacctaca accagctgtt cgaggaaaac cccatcaacg ccagcggcgt ggacgccaag
     5821 gccatcctgt ctgccagact gagcaagagc agacggctgg aaaatctgat cgcccagctg
     5881 cccggcgaga agaagaatgg cctgttcgga aacctgattg ccctgagcct gggcctgacc
     5941 cccaacttca agagcaactt cgacctggcc gaggatgcca aactgcagct gagcaaggac
     6001 acctacgacg acgacctgga caacctgctg gcccagatcg gcgaccagta cgccgacctg
     6061 tttctggccg ccaagaacct gtccgacgcc atcctgctga gcgacatcct gagagtgaac
     6121 accgagatca ccaaggcccc cctgagcgcc tctatgatca agagatacga cgagcaccac
     6181 caggacctga ccctgctgaa agctctcgtg cggcagcagc tgcctgagaa gtacaaagag
     6241 attttcttcg accagagcaa gaacggctac gccggctaca ttgacggcgg agccagccag
     6301 gaagagttct acaagttcat caagcccatc ctggaaaaga tggacggcac cgaggaactg
     6361 ctcgtgaagc tgaacagaga ggacctgctg cggaagcagc ggaccttcga caacggcagc
     6421 atcccccacc agatccacct gggagagctg cacgccattc tgcggcggca ggaagatttt
     6481 tacccattcc tgaaggacaa ccgggaaaag atcgagaaga tcctgacctt ccgcatcccc
     6541 tactacgtgg gccctctggc caggggaaac agcagattcg cctggatgac cagaaagagc
     6601 gaggaaacca tcaccccctg gaacttcgag gaagtggtgg acaagggcgc ttccgcccag
     6661 agcttcatcg agcggatgac caacttcgat aagaacctgc ccaacgagaa ggtgctgccc
     6721 aagcacagcc tgctgtacga gtacttcacc gtgtataacg agctgaccaa agtgaaatac
     6781 gtgaccgagg gaatgagaaa gcccgccttc ctgagcggcg agcagaaaaa ggccatcgtg
     6841 gacctgctgt tcaagaccaa ccggaaagtg accgtgaagc agctgaaaga ggactacttc
     6901 aagaaaatcg agtgcttcga ctccgtggaa atctccggcg tggaagatcg gttcaacgcc
     6961 tccctgggca cataccacga tctgctgaaa attatcaagg acaaggactt cctggacaat
     7021 gaggaaaacg aggacattct ggaagatatc gtgctgaccc tgacactgtt tgaggacaga
     7081 gagatgatcg aggaacggct gaaaacctat gcccacctgt tcgacgacaa agtgatgaag
     7141 cagctgaagc ggcggagata caccggctgg ggcaggctga gccggaagct gatcaacggc
     7201 atccgggaca agcagtccgg caagacaatc ctggatttcc tgaagtccga cggcttcgcc
     7261 aacagaaact tcatgcagct gatccacgac gacagcctga cctttaaaga ggacatccag
     7321 aaagcccagg tgtccggcca gggcgatagc ctgcacgagc acattgccaa tctggccggc
     7381 agccccgcca ttaagaaggg catcctgcag acagtgaagg tggtggacga gctcgtgaaa
     7441 gtgatgggcc ggcacaagcc cgagaacatc gtgatcgaaa tggccagaga gaaccagacc
     7501 acccagaagg gacagaagaa cagccgcgag agaatgaagc ggatcgaaga gggcatcaaa
     7561 gagctgggca gccagatcct gaaagaacac cccgtggaaa acacccagct gcagaacgag
     7621 aagctgtacc tgtactacct gcagaatggg cgggatatgt acgtggacca ggaactggac
     7681 atcaaccggc tgtccgacta cgatgtggac catatcgtgc ctcagagctt tctgaaggac
     7741 gactccatcg acaacaaggt gctgaccaga agcgacaaga accggggcaa gagcgacaac
     7801 gtgccctccg aagaggtcgt gaagaagatg aagaactact ggcggcagct gctgaacgcc
     7861 aagctgatta cccagagaaa gttcgacaat ctgaccaagg ccgagagagg cggcctgagc
     7921 gaactggata aggccggctt catcaagaga cagctggtgg aaacccggca gatcacaaag
     7981 cacgtggcac agatcctgga ctcccggatg aacactaagt acgacgagaa tgacaagctg
     8041 atccgggaag tgaaagtgat caccctgaag tccaagctgg tgtccgattt ccggaaggat
     8101 ttccagtttt acaaagtgcg cgagatcaac aactaccacc acgcccacga cgcctacctg
     8161 aacgccgtcg tgggaaccgc cctgatcaaa aagtacccta agctggaaag cgagttcgtg
     8221 tacggcgact acaaggtgta cgacgtgcgg aagatgatcg ccaagagcga gcaggaaatc
     8281 ggcaaggcta ccgccaagta cttcttctac agcaacatca tgaacttttt caagaccgag
     8341 attaccctgg ccaacggcga gatccggaag cggcctctga tcgagacaaa cggcgaaacc
     8401 ggggagatcg tgtgggataa gggccgggat tttgccaccg tgcggaaagt gctgagcatg
     8461 ccccaagtga atatcgtgaa aaagaccgag gtgcagacag gcggcttcag caaagagtct
     8521 atcctgccca agaggaacag cgataagctg atcgccagaa agaaggactg ggaccctaag
     8581 aagtacggcg gcttcgacag ccccaccgtg gcctattctg tgctggtggt ggccaaagtg
     8641 gaaaagggca agtccaagaa actgaagagt gtgaaagagc tgctggggat caccatcatg
     8701 gaaagaagca gcttcgagaa gaatcccatc gactttctgg aagccaaggg ctacaaagaa
     8761 gtgaaaaagg acctgatcat caagctgcct aagtactccc tgttcgagct ggaaaacggc
     8821 cggaagagaa tgctggcctc tgccggcgaa ctgcagaagg gaaacgaact ggccctgccc
     8881 tccaaatatg tgaacttcct gtacctggcc agccactatg agaagctgaa gggctccccc
     8941 gaggataatg agcagaaaca gctgtttgtg gaacagcaca agcactacct ggacgagatc
     9001 atcgagcaga tcagcgagtt ctccaagaga gtgatcctgg ccgacgctaa tctggacaaa
     9061 gtgctgtccg cctacaacaa gcaccgggat aagcccatca gagagcaggc cgagaatatc
     9121 atccacctgt ttaccctgac caatctggga gcccctgccg ccttcaagta ctttgacacc
     9181 accatcgacc ggaagaggta caccagcacc aaagaggtgc tggacgccac cctgatccac
     9241 cagagcatca ccggcctgta cgagacacgg atcgacctgt ctcagctggg aggcgacaag
     9301 cgacctgccg ccacaaagaa ggctggacag gctaagaaga agaaagatta caaagacgat
     9361 gacgataagg gatccggcgc aacaaacttc tctctgctga aacaagccgg agatgtcgaa
     9421 gagaatcctg gaccgaccga gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc
     9481 cccagggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac gcgccacacc
     9541 gtcgatccgg accgccacat cgagcgggtc accgagctgc aagaactctt cctcacgcgc
     9601 gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg
     9661 accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc gcgcatggcc
     9721 gagttgagcg gttcccggct ggccgcgcag caacagatgg aaggcctcct ggcgccgcac
     9781 cggcccaagg agcccgcgtg gttcctggcc accgtcggag tctcgcccga ccaccagggc
     9841 aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg cgccggggtg
     9901 cccgccttcc tggagacctc cgcgccccgc aacctcccct tctacgagcg gctcggcttc
     9961 accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag
    10021 cccggtgcct gaacgcgtta agtcgacaat caacctctgg attacaaaat ttgtgaaaga
    10081 ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg
    10141 cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc
    10201 tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc
    10261 actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt
    10321 tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt
    10381 gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg
    10441 aaatcatcgt cctttccttg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg
    10501 tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg
    10561 ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg gatctccctt
    10621 tgggccgcct ccccgcgtcg actttaagac caatgactta caaggcagct gtagatctta
    10681 gccacttttt aaaagaaaag gggggactgg aagggctaat tcactcccaa cgaagacaag
    10741 atctgctttt tgcttgtact gggtctctct ggttagacca gatctgagcc tgggagctct
    10801 ctggctaact agggaaccca ctgcttaagc ctcaataaag cttgccttga gtgcttcaag
    10861 tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag atccctcaga cccttttagt
    10921 cagtgtggaa aatctctagc agggcccgtt taaacccgct gatcagcctc gactgtgcct
    10981 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt
    11041 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg
    11101 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac
    11161 aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc
    11221 tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
    11281 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
    11341 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
    11401 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag
    11461 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
    11521 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
    11581 tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
    11641 gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt
    11701 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
    11761 cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
    11821 atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc
    11881 cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg
    11941 ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc
    12001 taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct gatcagcacg
    12061 tgttgacaat taatcatcgg catagtatat cggcatagta taatacgaca aggtgaggaa
    12121 ctaaaccatg gccaagttga ccagtgccgt tccggtgctc accgcgcgcg acgtcgccgg
    12181 agcggtcgag ttctggaccg accggctcgg gttctcccgg gacttcgtgg aggacgactt
    12241 cgccggtgtg gtccgggacg acgtgaccct gttcatcagc gcggtccagg accaggtggt
    12301 gccggacaac accctggcct gggtgtgggt gcgcggcctg gacgagctgt acgccgagtg
    12361 gtcggaggtc gtgtccacga acttccggga cgcctccggg ccggccatga ccgagatcgg
    12421 cgagcagccg tgggggcggg agttcgccct gcgcgacccg gccggcaact gcgtgcactt
    12481 cgtggccgag gagcaggact gacacgtgct acgagatttc gattccaccg ccgccttcta
    12541 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
    12601 ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta
    12661 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
    12721 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag
    12781 ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
    12841 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
    12901 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
    12961 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gta
//
