LOCUS       sequence-319420-       13364 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Multiplex Guide RNAs to generate Nr4a1 knockout by CRISPR.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13364)
  AUTHORS   Zhang Y, Federation AJ, Kim S, O'Keefe JP, Lun M, Xiang D, Brown JD,
            Steinhauser ML
  TITLE     Targeting nuclear receptor NR4A1-dependent adipocyte progenitor 
            quiescence promotes metabolic adaptation to obesity.
  JOURNAL   J Clin Invest. 2018 Nov 1;128(11):4898-4911. doi: 10.1172/JCI98353. 
            Epub 2018 Oct 2.
  PUBMED    30277475
REFERENCE   2  (bases 1 to 13364)
  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: "10.1172/JCI98353"; 
            journalName: "J Clin Invest"; date: "2018-11-1- 1"; volume: "128"; 
            issue: "11"; pages: "4898-4911"
COMMENT     Sequence Label: lentiCRISPR v2-Nr4a1 double gRNAs
FEATURES             Location/Qualifiers
     source          1..13364
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(47..66)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        238..617
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..817
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     772..792
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             835..1015
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1062..1187
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1680..1913
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2098..2142
                     /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    2440..2557
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2608..2856
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        2878..2953
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        2966..3214
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        3235..3310
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        3366..3577
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      3596..3605
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3602..7705
                     /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             3602..7705
                     /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             7706..7753
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7754..7777
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             7787..7843
                     /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             7844..8440
                     /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     8337..8357
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    8456..9044
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8509..8529)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8927..8938)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             9116..9349
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(9375..9392)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9381..9605
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      9651..10079
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9738..9757)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9948..9969
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10088..10108)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10093..10422
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      10273..10408
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     10335..10354
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        10470..10517
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             10536..10910
                     /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    11040..11161
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11077..11096)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11131..11150
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(11210..11226)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11210..11226)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11223..11245)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    11234..11250
                     /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(11258..11264,11265..11282,11283..11288))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11303..11324
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(11441..11458)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11612..12200)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(11692..11711)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12371..13162,13163..13231))
                     /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     12994..13013
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(13232..13336)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
       61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
      121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
      181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
      241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
      301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
      361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
      661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
      721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
      781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct
      841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
      901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
      961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc
     1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc
     1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa
     1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg
     1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata
     1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc
     1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga
     1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc
     1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca
     1501 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg
     1561 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga
     1621 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata
     1681 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg
     1741 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg
     1801 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag
     1861 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt
     1921 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt
     1981 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt
     2041 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag
     2101 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata
     2161 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta
     2221 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta
     2281 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa
     2341 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt
     2401 gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat
     2461 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa
     2521 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag
     2581 agatccagtt tggttaatta aggtaccgag ggcctatttc ccatgattcc ttcatatttg
     2641 catatacgat acaaggctgt tagagagata attagaatta atttgactgt aaacacaaag
     2701 atattagtac aaaatacgtg acgtagaaag taataatttc ttgggtagtt tgcagtttta
     2761 aaattatgtt ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta tttcgatttc
     2821 ttggctttat atatcttgtg gaaaggacga aacaccgccg ggtagcagcc gtacaccgtt
     2881 ttagagctag aaatagcaag ttaaaataag gctagtccgt tatcaacttg aaaaagtggc
     2941 accgagtcgg tgcttttttg aattcgaggg cctatttccc atgattcctt catatttgca
     3001 tatacgatac aaggctgtta gagagataat tagaattaat ttgactgtaa acacaaagat
     3061 attagtacaa aatacgtgac gtagaaagta ataatttctt gggtagtttg cagttttaaa
     3121 attatgtttt aaaatggact atcatatgct taccgtaact tgaaagtatt tcgatttctt
     3181 ggctttatat atcttgtgga aaggacgaaa caccgtccaa gtgtgcccgg atgagtttta
     3241 gagctagaaa tagcaagtta aaataaggct agtccgttat caacttgaaa aagtggcacc
     3301 gagtcggtgc ttttttgcta gctaggtctt gaaaggagtg ggaattggct ccggtgcccg
     3361 tcagtgggca gagcgcacat cgcccacagt ccccgagaag ttggggggag gggtcggcaa
     3421 ttgatccggt gcctagagaa ggtggcgcgg ggtaaactgg gaaagtgatg tcgtgtactg
     3481 gctccgcctt tttcccgagg gtgggggaga accgtatata agtgcagtag tcgccgtgaa
     3541 cgttcttttt cgcaacgggt ttgccgccag aacacaggac cggttctaga gcgctgccac
     3601 catggacaag aagtacagca tcggcctgga catcggcacc aactctgtgg gctgggccgt
     3661 gatcaccgac gagtacaagg tgcccagcaa gaaattcaag gtgctgggca acaccgaccg
     3721 gcacagcatc aagaagaacc tgatcggagc cctgctgttc gacagcggcg aaacagccga
     3781 ggccacccgg ctgaagagaa ccgccagaag aagatacacc agacggaaga accggatctg
     3841 ctatctgcaa gagatcttca gcaacgagat ggccaaggtg gacgacagct tcttccacag
     3901 actggaagag tccttcctgg tggaagagga taagaagcac gagcggcacc ccatcttcgg
     3961 caacatcgtg gacgaggtgg cctaccacga gaagtacccc accatctacc acctgagaaa
     4021 gaaactggtg gacagcaccg acaaggccga cctgcggctg atctatctgg ccctggccca
     4081 catgatcaag ttccggggcc acttcctgat cgagggcgac ctgaaccccg acaacagcga
     4141 cgtggacaag ctgttcatcc agctggtgca gacctacaac cagctgttcg aggaaaaccc
     4201 catcaacgcc agcggcgtgg acgccaaggc catcctgtct gccagactga gcaagagcag
     4261 acggctggaa aatctgatcg cccagctgcc cggcgagaag aagaatggcc tgttcggaaa
     4321 cctgattgcc ctgagcctgg gcctgacccc caacttcaag agcaacttcg acctggccga
     4381 ggatgccaaa ctgcagctga gcaaggacac ctacgacgac gacctggaca acctgctggc
     4441 ccagatcggc gaccagtacg ccgacctgtt tctggccgcc aagaacctgt ccgacgccat
     4501 cctgctgagc gacatcctga gagtgaacac cgagatcacc aaggcccccc tgagcgcctc
     4561 tatgatcaag agatacgacg agcaccacca ggacctgacc ctgctgaaag ctctcgtgcg
     4621 gcagcagctg cctgagaagt acaaagagat tttcttcgac cagagcaaga acggctacgc
     4681 cggctacatt gacggcggag ccagccagga agagttctac aagttcatca agcccatcct
     4741 ggaaaagatg gacggcaccg aggaactgct cgtgaagctg aacagagagg acctgctgcg
     4801 gaagcagcgg accttcgaca acggcagcat cccccaccag atccacctgg gagagctgca
     4861 cgccattctg cggcggcagg aagattttta cccattcctg aaggacaacc gggaaaagat
     4921 cgagaagatc ctgaccttcc gcatccccta ctacgtgggc cctctggcca ggggaaacag
     4981 cagattcgcc tggatgacca gaaagagcga ggaaaccatc accccctgga acttcgagga
     5041 agtggtggac aagggcgctt ccgcccagag cttcatcgag cggatgacca acttcgataa
     5101 gaacctgccc aacgagaagg tgctgcccaa gcacagcctg ctgtacgagt acttcaccgt
     5161 gtataacgag ctgaccaaag tgaaatacgt gaccgaggga atgagaaagc ccgccttcct
     5221 gagcggcgag cagaaaaagg ccatcgtgga cctgctgttc aagaccaacc ggaaagtgac
     5281 cgtgaagcag ctgaaagagg actacttcaa gaaaatcgag tgcttcgact ccgtggaaat
     5341 ctccggcgtg gaagatcggt tcaacgcctc cctgggcaca taccacgatc tgctgaaaat
     5401 tatcaaggac aaggacttcc tggacaatga ggaaaacgag gacattctgg aagatatcgt
     5461 gctgaccctg acactgtttg aggacagaga gatgatcgag gaacggctga aaacctatgc
     5521 ccacctgttc gacgacaaag tgatgaagca gctgaagcgg cggagataca ccggctgggg
     5581 caggctgagc cggaagctga tcaacggcat ccgggacaag cagtccggca agacaatcct
     5641 ggatttcctg aagtccgacg gcttcgccaa cagaaacttc atgcagctga tccacgacga
     5701 cagcctgacc tttaaagagg acatccagaa agcccaggtg tccggccagg gcgatagcct
     5761 gcacgagcac attgccaatc tggccggcag ccccgccatt aagaagggca tcctgcagac
     5821 agtgaaggtg gtggacgagc tcgtgaaagt gatgggccgg cacaagcccg agaacatcgt
     5881 gatcgaaatg gccagagaga accagaccac ccagaaggga cagaagaaca gccgcgagag
     5941 aatgaagcgg atcgaagagg gcatcaaaga gctgggcagc cagatcctga aagaacaccc
     6001 cgtggaaaac acccagctgc agaacgagaa gctgtacctg tactacctgc agaatgggcg
     6061 ggatatgtac gtggaccagg aactggacat caaccggctg tccgactacg atgtggacca
     6121 tatcgtgcct cagagctttc tgaaggacga ctccatcgac aacaaggtgc tgaccagaag
     6181 cgacaagaac cggggcaaga gcgacaacgt gccctccgaa gaggtcgtga agaagatgaa
     6241 gaactactgg cggcagctgc tgaacgccaa gctgattacc cagagaaagt tcgacaatct
     6301 gaccaaggcc gagagaggcg gcctgagcga actggataag gccggcttca tcaagagaca
     6361 gctggtggaa acccggcaga tcacaaagca cgtggcacag atcctggact cccggatgaa
     6421 cactaagtac gacgagaatg acaagctgat ccgggaagtg aaagtgatca ccctgaagtc
     6481 caagctggtg tccgatttcc ggaaggattt ccagttttac aaagtgcgcg agatcaacaa
     6541 ctaccaccac gcccacgacg cctacctgaa cgccgtcgtg ggaaccgccc tgatcaaaaa
     6601 gtaccctaag ctggaaagcg agttcgtgta cggcgactac aaggtgtacg acgtgcggaa
     6661 gatgatcgcc aagagcgagc aggaaatcgg caaggctacc gccaagtact tcttctacag
     6721 caacatcatg aactttttca agaccgagat taccctggcc aacggcgaga tccggaagcg
     6781 gcctctgatc gagacaaacg gcgaaaccgg ggagatcgtg tgggataagg gccgggattt
     6841 tgccaccgtg cggaaagtgc tgagcatgcc ccaagtgaat atcgtgaaaa agaccgaggt
     6901 gcagacaggc ggcttcagca aagagtctat cctgcccaag aggaacagcg ataagctgat
     6961 cgccagaaag aaggactggg accctaagaa gtacggcggc ttcgacagcc ccaccgtggc
     7021 ctattctgtg ctggtggtgg ccaaagtgga aaagggcaag tccaagaaac tgaagagtgt
     7081 gaaagagctg ctggggatca ccatcatgga aagaagcagc ttcgagaaga atcccatcga
     7141 ctttctggaa gccaagggct acaaagaagt gaaaaaggac ctgatcatca agctgcctaa
     7201 gtactccctg ttcgagctgg aaaacggccg gaagagaatg ctggcctctg ccggcgaact
     7261 gcagaaggga aacgaactgg ccctgccctc caaatatgtg aacttcctgt acctggccag
     7321 ccactatgag aagctgaagg gctcccccga ggataatgag cagaaacagc tgtttgtgga
     7381 acagcacaag cactacctgg acgagatcat cgagcagatc agcgagttct ccaagagagt
     7441 gatcctggcc gacgctaatc tggacaaagt gctgtccgcc tacaacaagc accgggataa
     7501 gcccatcaga gagcaggccg agaatatcat ccacctgttt accctgacca atctgggagc
     7561 ccctgccgcc ttcaagtact ttgacaccac catcgaccgg aagaggtaca ccagcaccaa
     7621 agaggtgctg gacgccaccc tgatccacca gagcatcacc ggcctgtacg agacacggat
     7681 cgacctgtct cagctgggag gcgacaagcg acctgccgcc acaaagaagg ctggacaggc
     7741 taagaagaag aaagattaca aagacgatga cgataaggga tccggcgcaa caaacttctc
     7801 tctgctgaaa caagccggag atgtcgaaga gaatcctgga ccgaccgagt acaagcccac
     7861 ggtgcgcctc gccacccgcg acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt
     7921 cgccgactac cccgccacgc gccacaccgt cgatccggac cgccacatcg agcgggtcac
     7981 cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc
     8041 ggacgacggc gccgcggtgg cggtctggac cacgccggag agcgtcgaag cgggggcggt
     8101 gttcgccgag atcggcccgc gcatggccga gttgagcggt tcccggctgg ccgcgcagca
     8161 acagatggaa ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt tcctggccac
     8221 cgtcggagtc tcgcccgacc accagggcaa gggtctgggc agcgccgtcg tgctccccgg
     8281 agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg gagacctccg cgccccgcaa
     8341 cctccccttc tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg
     8401 accgcgcacc tggtgcatga cccgcaagcc cggtgcctga acgcgttaag tcgacaatca
     8461 acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt
     8521 tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc
     8581 tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc
     8641 cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg
     8701 gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc
     8761 cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg
     8821 cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc tgctcgcctg
     8881 tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc
     8941 agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct
     9001 tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcgtcgac tttaagacca
     9061 atgacttaca aggcagctgt agatcttagc cactttttaa aagaaaaggg gggactggaa
     9121 gggctaattc actcccaacg aagacaagat ctgctttttg cttgtactgg gtctctctgg
     9181 ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct
     9241 caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt
     9301 aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag ggcccgttta
     9361 aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
     9421 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
     9481 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca
     9541 ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc
     9601 tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg
     9661 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     9721 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     9781 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     9841 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
     9901 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     9961 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
    10021 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta
    10081 attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
    10141 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
    10201 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
    10261 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
    10321 tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag
    10381 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt
    10441 atatccattt tcggatctga tcagcacgtg ttgacaatta atcatcggca tagtatatcg
    10501 gcatagtata atacgacaag gtgaggaact aaaccatggc caagttgacc agtgccgttc
    10561 cggtgctcac cgcgcgcgac gtcgccggag cggtcgagtt ctggaccgac cggctcgggt
    10621 tctcccggga cttcgtggag gacgacttcg ccggtgtggt ccgggacgac gtgaccctgt
    10681 tcatcagcgc ggtccaggac caggtggtgc cggacaacac cctggcctgg gtgtgggtgc
    10741 gcggcctgga cgagctgtac gccgagtggt cggaggtcgt gtccacgaac ttccgggacg
    10801 cctccgggcc ggccatgacc gagatcggcg agcagccgtg ggggcgggag ttcgccctgc
    10861 gcgacccggc cggcaactgc gtgcacttcg tggccgagga gcaggactga cacgtgctac
    10921 gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg
    10981 acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca
    11041 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
    11101 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
    11161 atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt
    11221 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
    11281 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
    11341 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
    11401 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
    11461 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
    11521 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
    11581 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
    11641 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
    11701 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
    11761 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
    11821 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
    11881 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
    11941 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
    12001 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
    12061 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
    12121 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
    12181 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
    12241 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
    12301 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
    12361 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
    12421 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
    12481 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
    12541 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
    12601 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
    12661 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
    12721 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
    12781 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
    12841 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
    12901 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
    12961 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
    13021 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
    13081 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
    13141 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
    13201 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
    13261 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
    13321 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgac
//
