LOCUS       sequence-178374-       12106 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  for SaCas9 knockout, lentiviral expression of S. aureus Cas9 
            (SauCas9) and gRNA scaffold.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12106)
  AUTHORS   Najm FJ, Strand C, Donovan KF, Hegde M, Sanson KR, Vaimberg EW, 
            Sullender ME, Hartenian E, Kalani Z, Fusi N, Listgarten J, Younger 
            ST, Bernstein BE, Root DE, Doench JG
  TITLE     Orthologous CRISPR-Cas9 enzymes for combinatorial genetic screens.
  JOURNAL   Nat Biotechnol. 2018 Feb;36(2):179-189. doi: 10.1038/nbt.4048. Epub 
            2017 Dec 18.
  PUBMED    29251726
REFERENCE   2  (bases 1 to 12106)
  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/nbt.4048"; 
            journalName: "Nat Biotechnol"; date: "2018-02"; volume: "36"; issue:
            "2"; pages: "179-189"
COMMENT     Sequence Label: pXPR_206
FEATURES             Location/Qualifiers
     source          1..12106
                     /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)"
     primer_bind     2608..2628
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2779..2798
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2881..2956
                     /label=Sa gRNA scaffold
                     /note="guide RNA scaffold for the Staphylococcus aureus 
                     CRISPR/Cas9 system"
     promoter        3018..3229
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      3237..3246
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3249..3269
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3294..6449
                     /codon_start=1
                     /product="Cas9 endonuclease from the Staphylococcus aureus 
                     Type II CRISPR/Cas system"
                     /label=SaCas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="KRNYILGLDIGITSVGYGIIDYETRDVIDAGVRLFKEANVENNEG
                     RRSKRGARRLKRRRRHRIQRVKKLLFDYNLLTDHSELSGINPYEARVKGLSQKLSEEEF
                     SAALLHLAKRRGVHNVNEVEEDTGNELSTKEQISRNSKALEEKYVAELQLERLKKDGEV
                     RGSINRFKTSDYVKEAKQLLKVQKAYHQLDQSFIDTYIDLLETRRTYYEGPGEGSPFGW
                     KDIKEWYEMLMGHCTYFPEELRSVKYAYNADLYNALNDLNNLVITRDENEKLEYYEKFQ
                     IIENVFKQKKKPTLKQIAKEILVNEEDIKGYRVTSTGKPEFTNLKVYHDIKDITARKEI
                     IENAELLDQIAKILTIYQSSEDIQEELTNLNSELTQEEIEQISNLKGYTGTHNLSLKAI
                     NLILDELWHTNDNQIAIFNRLKLVPKKVDLSQQKEIPTTLVDDFILSPVVKRSFIQSIK
                     VINAIIKKYGLPNDIIIELAREKNSKDAQKMINEMQKRNRQTNERIEEIIRTTGKENAK
                     YLIEKIKLHDMQEGKCLYSLEAIPLEDLLNNPFNYEVDHIIPRSVSFDNSFNNKVLVKQ
                     EENSKKGNRTPFQYLSSSDSKISYETFKKHILNLAKGKGRISKTKKEYLLEERDINRFS
                     VQKDFINRNLVDTRYATRGLMNLLRSYFRVNNLDVKVKSINGGFTSFLRRKWKFKKERN
                     KGYKHHAEDALIIANADFIFKEWKKLDKAKKVMENQMFEEKQAESMPEIETEQEYKEIF
                     ITPHQIKHIKDFKDYKYSHRVDKKPNRELINDTLYSTRKDDKGNTLIVNNLNGLYDKDN
                     DKLKKLINKSPEKLLMYHHDPQTYQKLKLIMEQYGDEKNPLYKYYEETGNYLTKYSKKD
                     NGPVIKKIKYYGNKLNAHLDITDDYPNSRNKVVKLSLKPYRFDVYLDNGVYKFVTVKNL
                     DVIKKENYYEVNSKCYEEAKKLKKISNQAEFIASFYNNDLIKINGELYRVIGVNNDLLN
                     RIEVNMIDITYREYLENMNDKRPPRIIKTIASKTQSIKKYSTDILGNLYEVKSKKHPQI
                     IKKG"
     CDS             6450..6497
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             6507..6563
                     /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"
     misc_feature    7192..7780
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7245..7265)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(7663..7674)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             7858..8091
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(8117..8134)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    8123..8347
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      8393..8821
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8480..8499)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     8690..8711
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(8830..8850)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        8835..9164
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      9015..9150
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     9077..9096
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        9212..9259
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             9278..9652
                     /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    9782..9903
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(9819..9838)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     9873..9892
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(9952..9968)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9952..9968)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(9965..9987)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    9976..9992
                     /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(10000..10006,10007..10024,10025..10030))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    10045..10066
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(10183..10200)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(10354..10942)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(10434..10453)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(11113..11904,11905..11973))
                     /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     11736..11755
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(11974..12078)
                     /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 aacaccggag acgaacctag catcgtctct
     2881 gtttaagtac tctggaaaca gaatctactt aaacaaggca aaatgccgtg tttatctcgt
     2941 caacttgttg gcgagatttt ttgaattcgc tagctaggtc ttgaaaggag tgggaattgg
     3001 ctccggtgcc cgtcagtggg cagagcgcac atcgcccaca gtccccgaga agttgggggg
     3061 aggggtcggc aattgatccg gtgcctagag aaggtggcgc ggggtaaact gggaaagtga
     3121 tgtcgtgtac tggctccgcc tttttcccga gggtggggga gaaccgtata taagtgcagt
     3181 agtcgccgtg aacgttcttt ttcgcaacgg gtttgccgcc agaacacagg accggtgcca
     3241 ccatggcacc caagaagaag cgcaaagttg gcatccacgg cgtgcctgct gccaagcgca
     3301 actacatcct gggcctggac attggcatca cctcagttgg ctatggcatc atcgactacg
     3361 agacccggga cgtcatcgac gctggcgtcc gcctcttcaa ggaggccaac gtcgagaaca
     3421 acgagggccg gcggtccaag cgcggggccc gccggctcaa gcgccgccgc cgccaccgca
     3481 tccagcgcgt gaaaaagttg ctcttcgact acaacttgct caccgaccac tccgagctgt
     3541 ctggcatcaa cccgtacgag gcccgggtca agggcctctc gcaaaagttg tccgaggagg
     3601 agttctctgc tgccctgctg cacctggcca agcgccgcgg ggtccacaac gtcaacgaag
     3661 ttgaggagga caccggcaac gaactttcca ccaaggagca gatctcccgc aactccaagg
     3721 ccctggagga gaagtacgtc gcggagctcc aacttgagcg cctcaagaag gacggcgagg
     3781 tgcgcggctc catcaaccgc ttcaagacct ccgactacgt caaggaggcc aagcagctgc
     3841 tcaaggtcca gaaggcctac caccaacttg accagtcctt catcgacacc tacatcgacc
     3901 tgctggagac ccgccgcacc tactacgagg ggcctggcga gggctctccc ttcggctgga
     3961 aggacatcaa ggagtggtac gagatgctca tgggccactg cacctacttc ccggaggagc
     4021 tgcgctcggt caagtacgcc tacaacgccg acctctacaa cgcgctcaac gacctcaaca
     4081 acttggtcat cacccgcgac gagaacgaaa agttggagta ctacgaaaag ttccagatca
     4141 tcgaaaacgt cttcaagcag aagaagaagc caactttgaa gcagatcgcc aaggagatcc
     4201 tggtcaacga ggaggacatc aagggctacc gggtcacctc caccggcaag cccgagttca
     4261 ccaacttaaa ggtctaccat gacatcaagg acatcaccgc ccgcaaggag atcatcgaga
     4321 acgcggagct gctggaccag atcgccaaga tcctcaccat ctaccaaagt tccgaggaca
     4381 tccaggagga gctgaccaac ttaaactcgg agcttaccca ggaggagatc gagcagatct
     4441 ccaacttaaa gggctacact ggcacccaca acttgtccct caaggccatc aacttgatcc
     4501 tggacgaact ttggcatacc aacgacaacc agatcgccat cttcaaccga ctaaagttgg
     4561 tgcccaagaa agttgacctg tcccaacaga aggagatccc aactaccctg gtcgacgact
     4621 tcatcctctc gcccgtggtc aagcgcagct tcatccagtc catcaaggtc atcaacgcca
     4681 tcatcaagaa gtacggcctg cccaacgaca tcatcatcga gttggcacgc gagaagaact
     4741 ccaaggatgc ccagaagatg atcaacgaga tgcagaagcg caaccgccag accaacgagc
     4801 gcatcgagga gatcatccgc accaccggca aggagaacgc caagtacctg atcgagaaga
     4861 taaagttgca tgacatgcag gagggcaagt gcctctactc cctggaggcc atccccctcg
     4921 aggacctgct caacaacccc ttcaactacg aagttgacca catcatcccg cgctcggtgt
     4981 ccttcgacaa cagcttcaac aacaaggtcc tggtcaagca ggaggagaac tccaagaagg
     5041 gcaaccggac ccccttccag tacctgtcct cctcggactc caagatctcc tacgaaactt
     5101 tcaagaagca catcctcaac ttggcaaagg gcaagggccg catctccaag accaagaagg
     5161 agtacctgct ggaggagcgc gacatcaacc gcttctcggt gcagaaggac ttcatcaacc
     5221 gcaacctggt ggacacccgc tacgcaaccc gtggcctcat gaacctgctg cgcagctact
     5281 tccgggtcaa caacttggac gtcaaagtta agtcgatcaa cggtggcttc accagcttcc
     5341 tgcgccgcaa gtggaagttc aagaaggagc gcaacaaggg ctacaagcac catgcggagg
     5401 acgccctcat catcgccaac gccgacttca tcttcaagga gtggaaaaag ttggacaagg
     5461 ccaagaaggt catggagaac cagatgttcg aggagaagca ggccgagtcc atgccggaga
     5521 tcgagacaga gcaggagtac aaggagatct tcatcacgcc gcaccagatc aagcacatca
     5581 aggacttcaa ggactacaag tactcccacc gcgtggacaa gaagcccaac cgcgagctca
     5641 tcaacgacac cctctactcc acccgcaagg acgacaaggg caacaccctc atcgtcaaca
     5701 acttaaacgg cctctacgac aaggacaacg acaagttgaa aaagttgatc aacaagagcc
     5761 ccgaaaagtt gctcatgtac caccatgacc cgcagacgta ccaaaagttg aagttgatca
     5821 tggagcagta cggtgacgag aagaacccgc tctacaagta ctacgaggag accggcaact
     5881 acctcaccaa gtactccaag aaggacaacg gcccggtcat caagaagatc aagtactacg
     5941 gcaacaagtt gaatgcccac ctggacatca ccgacgacta ccccaactcc cgcaacaaag
     6001 ttgtaaagtt gtcgctaaag ccgtaccgct tcgacgtcta cctggacaac ggcgtctaca
     6061 agttcgtgac cgtcaagaac ctggacgtca tcaagaagga gaactactac gaggtcaact
     6121 ccaagtgcta cgaggaggcc aaaaagttga agaagatctc caaccaggcc gagttcatcg
     6181 ccagcttcta caacaacgac ctcatcaaga tcaacggcga gctctaccgc gtcatcgggg
     6241 tcaacaacga cctgctgaac cgcatcgagg tcaacatgat cgacatcacc taccgcgagt
     6301 acctcgagaa catgaacgac aagcgcccgc ctcgcatcat caagaccatc gcctccaaga
     6361 cccagtccat caagaagtac tccaccgaca tccttggcaa cttgtacgag gtcaagagca
     6421 agaagcaccc tcagatcatc aagaagggca agcgccctgc tgccaccaag aaggctggcc
     6481 aggccaagaa gaagaaggga tccggggcca ccaacttttc cctgctgaag caggctggcg
     6541 acgtcgagga gaaccctggc cctggctccg gaaccgagta caagccaact gtccgcttgg
     6601 caacccgcga cgatgtgccc agggccgtcc gcacgctggc tgctgccttc gccgactacc
     6661 cagccacccg ccacaccgtg gacccggacc gccacatcga gcgcgtcacc gagctgcagg
     6721 agctcttcct cacccgagtt ggccttgaca ttggcaaggt ctgggtggcg gacgacggtg
     6781 ctgccgtggc cgtctggacc acccccgagt cggtcgaggc tggcgccgtc ttcgccgaga
     6841 tcgggccgcg catggccgag ctttcgggct cccggttggc tgcccaacag cagatggagg
     6901 ggctgttggc accccaccgc cccaaggagc cggcctggtt cctggccaca gttggtgtct
     6961 ctcccgacca ccagggcaag ggcctgggca gcgccgtggt gctgcctggg gtcgaggctg
     7021 ccgagcgcgc tggcgtccct gccttcctgg agacctctgc ccctcgcaac ttgcccttct
     7081 acgagcgcct gggcttcacc gtcaccgcgg acgtcgaggt gcccgagggg ccgcgcacct
     7141 ggtgcatgac ccgcaagcct ggggcatgaa tctagaacgc gttaagtcga caatcaacct
     7201 ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc tccttttacg
     7261 ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg tatggctttc
     7321 attttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt gtggcccgtt
     7381 gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg caacccccac tggttggggc
     7441 attgccacca cctgtcagct cctttccggg actttcgctt tccccctccc tattgccacg
     7501 gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct gttgggcact
     7561 gacaattccg tggtgttgtc ggggaaatca tcgtcctttc cttggctgct cgcctgtgtt
     7621 gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct caatccagcg
     7681 gaccttcctt cccgcggcct gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc
     7741 cctcagacga gtcggatctc cctttgggcc gcctccccgc agcgctgtcg actttaagac
     7801 caatgactta caaggcagct gtagatctta gccacttttt aaaagaaaag gggggactgg
     7861 aagggctaat tcactcccaa cgaagacaag atctgctttt tgcttgtact gggtctctct
     7921 ggttagacca gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc
     7981 ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg
     8041 gtaactagag atccctcaga cccttttagt cagtgtggaa aatctctagc agggcccgtt
     8101 taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc
     8161 tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat
     8221 gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg
     8281 caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc
     8341 tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc
     8401 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     8461 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     8521 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     8581 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
     8641 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     8701 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     8761 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     8821 taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
     8881 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     8941 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     9001 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     9061 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc
     9121 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt
     9181 gtatatccat tttcggatct gatcagcacg tgttgacaat taatcatcgg catagtatat
     9241 cggcatagta taatacgaca aggtgaggaa ctaaaccatg gccaagttga ccagtgccgt
     9301 tccggtgctc accgcgcgcg acgtcgccgg agcggtcgag ttctggaccg accggctcgg
     9361 gttctcccgg gacttcgtgg aggacgactt cgccggtgtg gtccgggacg acgtgaccct
     9421 gttcatcagc gcggtccagg accaggtggt gccggacaac accctggcct gggtgtgggt
     9481 gcgcggcctg gacgagctgt acgccgagtg gtcggaggtc gtgtccacga acttccggga
     9541 cgcctccggg ccggccatga ccgagatcgg cgagcagccg tgggggcggg agttcgccct
     9601 gcgcgacccg gccggcaact gcgtgcactt cgtggccgag gagcaggact gacacgtgct
     9661 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg
     9721 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc
     9781 caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     9841 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     9901 ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct
     9961 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
    10021 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
    10081 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
    10141 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
    10201 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
    10261 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
    10321 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
    10381 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
    10441 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
    10501 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
    10561 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
    10621 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
    10681 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
    10741 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
    10801 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
    10861 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
    10921 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
    10981 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
    11041 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
    11101 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
    11161 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
    11221 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
    11281 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
    11341 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
    11401 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
    11461 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
    11521 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
    11581 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
    11641 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
    11701 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
    11761 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
    11821 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
    11881 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
    11941 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
    12001 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
    12061 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgac
//
