LOCUS       sequence-384742-       13813 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses gRNA and VP64-dSaCas9-VP64 from lentiviral vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13813)
  AUTHORS   McCutcheon SR, Swartz AM, Brown MC, Barrera A, McRoberts Amador C, 
            Siklenka K, Humayun L, Isaacs JM, Reddy TE, Nair S, Antonia S, 
            Gersbach CA
  TITLE     Orthogonal CRISPR screens to identify transcriptional and epigenetic
            regulators of human CD8 T cell function.
  JOURNAL   bioRxiv. 2023 May 1:2023.05.01.538906. doi: 
            10.1101/2023.05.01.538906. Preprint.
  PUBMED    37205457
REFERENCE   2  (bases 1 to 13813)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv. 
            2023 May 1:2023.05.01.538906. doi: 10.1101/2023.05.01.538906. 
            Preprint."
COMMENT     Sequence Label: pLV hU6-gRNA hUbC-VP64-dSaCas9-VP64-T2A-Thy1.1
FEATURES             Location/Qualifiers
     source          1..13813
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(15..34)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        206..585
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        587..785
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     740..760
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             803..983
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1030..1155
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1648..1881
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2066..2110
                     /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    2408..2525
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     misc_RNA        complement(2608..2683)
                     /label=Sa gRNA scaffold
                     /note="guide RNA scaffold for the Staphylococcus aureus 
                     CRISPR/Cas9 system"
     primer_bind     complement(2758..2777)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     promoter        2970..4181
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4018..4037
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4219..4245
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             4246..4395
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     CDS             4426..4446
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7627..7674
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7705..7854
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     CDS             7879..7899
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7906..7959
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             7960..8448
                     /codon_start=1
                     /product="mouse GPI-anchored cell surface glycoprotein of 
                     the immunoglobulin superfamily"
                     /label=Thy1.1
                     /note="This allele of Thy1 is also known as CD90.1."
                     /translation="MNPAISVALLLSVLQVSRGQKVTSLTACLVNQNLRLDCRHENNTK
                     DNSIQHEFSLTREKRKHVLSGTLGIPEHTYRSRVTLSNQPYIKVLTLANFTTKDEGDYF
                     CELRVSGANPMSSNKSISVYRDKLVKCGGISLLVQNTSWMLLLLLSLSLLQALDFISL"
     misc_feature    8473..9061
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8526..8546)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8944..8955)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(9064..9080)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9065..9081)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             9586..9766
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(9792..9809)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9798..10022
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      10068..10496
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(10155..10174)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10365..10386
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10505..10525)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10510..10839
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      10690..10825
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     10752..10771
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        10887..10934
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             10953..11327
                     /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    11457..11578
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11494..11513)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11548..11567
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(11627..11643)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11627..11643)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11640..11662)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    11651..11667
                     /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(11675..11681,11682..11699,11700..11705))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11720..11741
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(11858..11875)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(12029..12617)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(12109..12128)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12788..13579,13580..13648))
                     /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     13411..13430
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(13649..13753)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta tctgctccct
       61 gcttgtgtgt tggaggtcgc tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa
      121 ggcttgaccg acaattgcat gaagaatctg cttagggtta ggcgttttgc gctgcttcgc
      181 gatgtacggg ccagatatac gcgttgacat tgattattga ctagttatta atagtaatca
      241 attacggggt cattagttca tagcccatat atggagttcc gcgttacata acttacggta
      301 aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat
      361 gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg
      421 taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac
      481 gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt
      541 cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg
      601 cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc
      661 attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt
      721 aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata
      781 agcagcgcgt tttgcctgta ctgggtctct ctggttagac cagatctgag cctgggagct
      841 ctctggctaa ctagggaacc cactgcttaa gcctcaataa agcttgcctt gagtgcttca
      901 agtagtgtgt gcccgtctgt tgtgtgactc tggtaactag agatccctca gaccctttta
      961 gtcagtgtgg aaaatctcta gcagtggcgc ccgaacaggg acttgaaagc gaaagggaaa
     1021 ccagaggagc tctctcgacg caggactcgg cttgctgaag cgcgcacggc aagaggcgag
     1081 gggcggcgac tggtgagtac gccaaaaatt ttgactagcg gaggctagaa ggagagagat
     1141 gggtgcgaga gcgtcagtat taagcggggg agaattagat cgcgatggga aaaaattcgg
     1201 ttaaggccag ggggaaagaa aaaatataaa ttaaaacata tagtatgggc aagcagggag
     1261 ctagaacgat tcgcagttaa tcctggcctg ttagaaacat cagaaggctg tagacaaata
     1321 ctgggacagc tacaaccatc ccttcagaca ggatcagaag aacttagatc attatataat
     1381 acagtagcaa ccctctattg tgtgcatcaa aggatagaga taaaagacac caaggaagct
     1441 ttagacaaga tagaggaaga gcaaaacaaa agtaagacca ccgcacagca agcggccgct
     1501 gatcttcaga cctggaggag gagatatgag ggacaattgg agaagtgaat tatataaata
     1561 taaagtagta aaaattgaac cattaggagt agcacccacc aaggcaaaga gaagagtggt
     1621 gcagagagaa aaaagagcag tgggaatagg agctttgttc cttgggttct tgggagcagc
     1681 aggaagcact atgggcgcag cgtcaatgac gctgacggta caggccagac aattattgtc
     1741 tggtatagtg cagcagcaga acaatttgct gagggctatt gaggcgcaac agcatctgtt
     1801 gcaactcaca gtctggggca tcaagcagct ccaggcaaga atcctggctg tggaaagata
     1861 cctaaaggat caacagctcc tggggatttg gggttgctct ggaaaactca tttgcaccac
     1921 tgctgtgcct tggaatgcta gttggagtaa taaatctctg gaacagattt ggaatcacac
     1981 gacctggatg gagtgggaca gagaaattaa caattacaca agcttaatac actccttaat
     2041 tgaagaatcg caaaaccagc aagaaaagaa tgaacaagaa ttattggaat tagataaatg
     2101 ggcaagtttg tggaattggt ttaacataac aaattggctg tggtatataa aattattcat
     2161 aatgatagta ggaggcttgg taggtttaag aatagttttt gctgtacttt ctatagtgaa
     2221 tagagttagg cagggatatt caccattatc gtttcagacc cacctcccaa ccccgagggg
     2281 acccgacagg cccgaaggaa tagaagaaga aggtggagag agagacagag acagatccat
     2341 tcgattagtg aacggatcgg cactgcgtgc gccaattctg cagacaaatg gcagtattca
     2401 tccacaattt taaaagaaaa ggggggattg gggggtacag tgcaggggaa agaatagtag
     2461 acataatagc aacagacata caaactaaag aattacaaaa acaaattaca aaaattcaaa
     2521 attttcgggt ttattacagg gacagcagag atccagtttg gttaatgaac aaaagctgga
     2581 gctccaccgc gggcggccgc aaaaaaatct cgccaacaag ttgacgagat aaacacggca
     2641 ttttgccttg ttttagtaga ttctgtttcc agagtactaa aacagagacg tccgtctccg
     2701 gtgtttcgtc ctttccacaa gatatataaa gccaagaaat cgaaatactt tcaagttacg
     2761 gtaagcatat gatagtccat tttaaaacat aattttaaaa ctgcaaacta cccaagaaat
     2821 tattactttc tacgtcacgt attttgtact aatatctttg tgtttacagt caaattaatt
     2881 ccaattatct ctctaacagc cttgtatcgt atatgcaaat atgaaggaat catgggaaat
     2941 aggcattaac ccgtgtcggc tccagatctg gcctccgcgc cgggttttgg cgcctcccgc
     3001 gggcgccccc ctcctcacgg cgagcgctgc cacgtcagac gaagggcgca gcgagcgtcc
     3061 tgatccttcc gcccggacgc tcaggacagc ggcccgctgc tcataagact cggccttaga
     3121 accccagtat cagcagaagg acattttagg acgggacttg ggtgactcta gggcactggt
     3181 tttctttcca gagagcggaa caggcgagga aaagtagtcc cttctcggcg attctgcgga
     3241 gggatctccg tggggcggtg aacgccgatg attatataag gacgcgccgg gtgtggcaca
     3301 gctagttccg tcgcagccgg gatttgggtc gcggttcttg tttgtggatc gctgtgatcg
     3361 tcacttggtg agtagcgggc tgctgggctg gccggggctt tcgtggccgc cgggccgctc
     3421 ggtgggacgg aagcgtgtgg agagaccgcc aagggctgta gtctgggtcc gcgagcaagg
     3481 ttgccctgaa ctgggggttg gggggagcgc agcaaaatgg cggctgttcc cgagtcttga
     3541 atggaagacg cttgtgaggc gggctgtgag gtcgttgaaa caaggtgggg ggcatggtgg
     3601 gcggcaagaa cccaaggtct tgaggccttc gctaatgcgg gaaagctctt attcgggtga
     3661 gatgggctgg ggcaccatct ggggaccctg acgtgaagtt tgtcactgac tggagaactc
     3721 ggtttgtcgt ctgttgcggg ggcggcagtt atggcggtgc cgttgggcag tgcacccgta
     3781 cctttgggag cgcgcgccct cgtcgtgtcg tgacgtcacc cgttctgttg gcttataatg
     3841 cagggtgggg ccacctgccg gtaggtgtgc ggtaggcttt tctccgtcgc aggacgcagg
     3901 gttcgggcct agggtaggct ctcctgaatc gacaggcgcc ggacctctgg tgaggggagg
     3961 gataagtgag gcgtcagttt ctttggtcgg ttttatgtac ctatcttctt aagtagctga
     4021 agctccggtt ttgaactatg cgctcggggt tggcgagtgt gttttgtgaa gttttttagg
     4081 caccttttga aatgtaatca tttgggtcaa tatgtaattt tcagtgttag actagtaaat
     4141 tgtccgctaa attctggccg tttttggctt ttttgttaga cgaagcttgg gctgcaggtc
     4201 gactctagag ccaccatgta cccatacgat gttccagatt acgctgacgc gctggacgat
     4261 tttgacctcg acatgctggg ttctgatgcc ctcgatgact ttgacctgga tatgttggga
     4321 agcgacgcat tggatgactt tgatctggac atgctcggct ccgatgctct ggacgatttc
     4381 gatctcgata tgttaattaa ctctaggagt tccggctcta tggccccaaa gaagaagcgg
     4441 aaggtcggta tccacggagt cccagcagcc aagcggaact acatcctggg cctggccatc
     4501 ggcatcacca gcgtgggcta cggcatcatc gactacgaga cacgggacgt gatcgatgcc
     4561 ggcgtgcggc tgttcaaaga ggccaacgtg gaaaacaacg agggcaggcg gagcaagaga
     4621 ggcgccagaa ggctgaagcg gcggaggcgg catagaatcc agagagtgaa gaagctgctg
     4681 ttcgactaca acctgctgac cgaccacagc gagctgagcg gcatcaaccc ctacgaggcc
     4741 agagtgaagg gcctgagcca gaagctgagc gaggaagagt tctctgccgc cctgctgcac
     4801 ctggccaaga gaagaggcgt gcacaacgtg aacgaggtgg aagaggacac cggcaacgag
     4861 ctgtccacca aagagcagat cagccggaac agcaaggccc tggaagagaa atacgtggcc
     4921 gaactgcagc tggaacggct gaagaaagac ggcgaagtgc ggggcagcat caacagattc
     4981 aagaccagcg actacgtgaa agaagccaaa cagctgctga aggtgcagaa ggcctaccac
     5041 cagctggacc agagcttcat cgacacctac atcgacctgc tggaaacccg gcggacctac
     5101 tatgagggac ctggcgaggg cagccccttc ggctggaagg acatcaaaga atggtacgag
     5161 atgctgatgg gccactgcac ctacttcccc gaggaactgc ggagcgtgaa gtacgcctac
     5221 aacgccgacc tgtacaacgc cctgaacgac ctgaacaatc tcgtgatcac cagggacgag
     5281 aacgagaagc tggaatatta cgagaagttc cagatcatcg agaacgtgtt caagcagaag
     5341 aagaagccca ccctgaagca gatcgccaaa gaaatcctcg tgaacgaaga ggatattaag
     5401 ggctacagag tgaccagcac cggcaagccc gagttcacca acctgaaggt gtaccacgac
     5461 atcaaggaca ttaccgcccg gaaagagatt attgagaacg ccgagctgct ggatcagatt
     5521 gccaagatcc tgaccatcta ccagagcagc gaggacatcc aggaagaact gaccaatctg
     5581 aactccgagc tgacccagga agagatcgag cagatctcta atctgaaggg ctataccggc
     5641 acccacaacc tgagcctgaa ggccatcaac ctgatcctgg acgagctgtg gcacaccaac
     5701 gacaaccaga tcgctatctt caaccggctg aagctggtgc ccaagaaggt ggacctgtcc
     5761 cagcagaaag agatccccac caccctggtg gacgacttca tcctgagccc cgtcgtgaag
     5821 agaagcttca tccagagcat caaagtgatc aacgccatca tcaagaagta cggcctgccc
     5881 aacgacatca ttatcgagct ggcccgcgag aagaactcca aggacgccca gaaaatgatc
     5941 aacgagatgc agaagcggaa ccggcagacc aacgagcgga tcgaggaaat catccggacc
     6001 accggcaaag agaacgccaa gtacctgatc gagaagatca agctgcacga catgcaggaa
     6061 ggcaagtgcc tgtacagcct ggaagccatc cctctggaag atctgctgaa caaccccttc
     6121 aactatgagg tggaccacat catccccaga agcgtgtcct tcgacaacag cttcaacaac
     6181 aaggtgctcg tgaagcagga agaagccagc aagaagggca accggacccc attccagtac
     6241 ctgagcagca gcgacagcaa gatcagctac gaaaccttca agaagcacat cctgaatctg
     6301 gccaagggca agggcagaat cagcaagacc aagaaagagt atctgctgga agaacgggac
     6361 atcaacaggt tctccgtgca gaaagacttc atcaaccgga acctggtgga taccagatac
     6421 gccaccagag gcctgatgaa cctgctgcgg agctacttca gagtgaacaa cctggacgtg
     6481 aaagtgaagt ccatcaatgg cggcttcacc agctttctgc ggcggaagtg gaagtttaag
     6541 aaagagcgga acaaggggta caagcaccac gccgaggacg ccctgatcat tgccaacgcc
     6601 gatttcatct tcaaagagtg gaagaaactg gacaaggcca aaaaagtgat ggaaaaccag
     6661 atgttcgagg aaaagcaggc cgagagcatg cccgagatcg aaaccgagca ggagtacaaa
     6721 gagatcttca tcacccccca ccagatcaag cacattaagg acttcaagga ctacaagtac
     6781 agccaccggg tggacaagaa gcctaataga gagctgatta acgacaccct gtactccacc
     6841 cggaaggacg acaagggcaa caccctgatc gtgaacaatc tgaacggcct gtacgacaag
     6901 gacaatgaca agctgaaaaa gctgatcaac aagagccccg aaaagctgct gatgtaccac
     6961 cacgaccccc agacctacca gaaactgaag ctgattatgg aacagtacgg cgacgagaag
     7021 aatcccctgt acaagtacta cgaggaaacc gggaactacc tgaccaagta ctccaaaaag
     7081 gacaacggcc ccgtgatcaa gaagattaag tattacggca acaaactgaa cgcccatctg
     7141 gacatcaccg acgactaccc caacagcaga aacaaggtcg tgaagctgtc cctgaagccc
     7201 tacagattcg acgtgtacct ggacaatggc gtgtacaagt tcgtgaccgt gaagaatctg
     7261 gatgtgatca aaaaagaaaa ctactacgaa gtgaatagca agtgctatga ggaagctaag
     7321 aagctgaaga agatcagcaa ccaggccgag tttatcgcct ccttctacaa caacgatctg
     7381 atcaagatca acggcgagct gtatagagtg atcggcgtga acaacgacct gctgaaccgg
     7441 atcgaagtga acatgatcga catcacctac cgcgagtacc tggaaaacat gaacgacaag
     7501 aggcccccca ggatcattaa gacaatcgcc tccaagaccc agagcattaa gaagtacagc
     7561 acagacattc tgggcaacct gtatgaagtg aaatctaaga agcaccctca gatcatcaaa
     7621 aagggcaaaa ggccggcggc cacgaaaaag gccggccagg caaaaaagaa aaagggatcc
     7681 gaggccagcg gttccggacg ggctgacgca ttggacgatt ttgatctgga tatgctggga
     7741 agtgacgccc tcgatgattt tgaccttgac atgcttggtt cggatgccct tgatgacttt
     7801 gacctcgaca tgctcggcag tgacgccctt gatgatttcg acctggacat gctgattaac
     7861 tctagaagtt ccggatctcc gaaaaagaaa cgcaaagttg ctagcgaggg cagaggaagt
     7921 cttctaacat gcggtgacgt ggaggagaat cccggcccta tgaacccagc catcagcgtc
     7981 gctctcctgc tctcagtctt gcaggtgtcc cgagggcaga aggtgaccag cctgacagcc
     8041 tgcctggtga accaaaacct tcgcctggac tgccgccatg agaataacac caaggataac
     8101 tccatccagc atgagttcag cctgacccga gagaagagga agcacgtgct ctcaggcacc
     8161 cttgggatac ccgagcacac gtaccgctcc cgcgtcaccc tctccaacca gccctatatc
     8221 aaggtcctta ccctagccaa cttcaccacc aaggatgagg gcgactactt ttgtgagctt
     8281 cgcgtttcgg gcgcgaatcc catgagctcc aataaaagta tcagtgtgta tagagacaag
     8341 ctggtcaagt gtggcggcat aagcctgctg gttcagaaca catcctggat gctgctgctg
     8401 ctgctttccc tctccctcct ccaagccctg gacttcattt ctctgtgaga attcgatatc
     8461 aagcttatcg ataatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt
     8521 aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct
     8581 attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt
     8641 tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac
     8701 gcaaccccca ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct
     8761 ttccccctcc ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca
     8821 ggggctcggc tgttgggcac tgacaattcc gtggtgttgt cggggaaatc atcgtccttt
     8881 ccttggctgc tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc
     8941 ccttcggccc tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct
     9001 cttccgcgtc ttcgccttcg ccctcagacg agtcggatct ccctttgggc cgcctccccg
     9061 catcgatacc gtcgacctcg agacctagaa aaacatggag caatcacaag tagcaataca
     9121 gcagctacca atgctgattg tgcctggcta gaagcacaag aggaggagga ggtgggtttt
     9181 ccagtcacac ctcaggtacc tttaagacca atgacttaca aggcagctgt agatcttagc
     9241 cactttttaa aagaaaaggg gggactggaa gggctaattc actcccaacg aagacaagat
     9301 atccttgatc tgtggatcta ccacacacaa ggctacttcc ctgattggca gaactacaca
     9361 ccagggccag ggatcagata tccactgacc tttggatggt gctacaagct agtaccagtt
     9421 gagcaagaga aggtagaaga agccaatgaa ggagagaaca cccgcttgtt acaccctgtg
     9481 agcctgcatg ggatggatga cccggagaga gaagtattag agtggaggtt tgacagccgc
     9541 ctagcatttc atcacatggc ccgagagctg catccggact gtactgggtc tctctggtta
     9601 gaccagatct gagcctggga gctctctggc taactaggga acccactgct taagcctcaa
     9661 taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac
     9721 tagagatccc tcagaccctt ttagtcagtg tggaaaatct ctagcagggc ccgtttaaac
     9781 ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc
     9841 cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga
     9901 aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga
     9961 cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat
    10021 ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag
    10081 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
    10141 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
    10201 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
    10261 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
    10321 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
    10381 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
    10441 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattaatt
    10501 ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt
    10561 atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca
    10621 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta
    10681 actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga
    10741 ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct attccagaag
    10801 tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg agcttgtata
    10861 tccattttcg gatctgatca gcacgtgttg acaattaatc atcggcatag tatatcggca
    10921 tagtataata cgacaaggtg aggaactaaa ccatggccaa gttgaccagt gccgttccgg
    10981 tgctcaccgc gcgcgacgtc gccggagcgg tcgagttctg gaccgaccgg ctcgggttct
    11041 cccgggactt cgtggaggac gacttcgccg gtgtggtccg ggacgacgtg accctgttca
    11101 tcagcgcggt ccaggaccag gtggtgccgg acaacaccct ggcctgggtg tgggtgcgcg
    11161 gcctggacga gctgtacgcc gagtggtcgg aggtcgtgtc cacgaacttc cgggacgcct
    11221 ccgggccggc catgaccgag atcggcgagc agccgtgggg gcgggagttc gccctgcgcg
    11281 acccggccgg caactgcgtg cacttcgtgg ccgaggagca ggactgacac gtgctacgag
    11341 atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg
    11401 ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact
    11461 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
    11521 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
    11581 atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc
    11641 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
    11701 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
    11761 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
    11821 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
    11881 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
    11941 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
    12001 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
    12061 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
    12121 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
    12181 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
    12241 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
    12301 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
    12361 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
    12421 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
    12481 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
    12541 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
    12601 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
    12661 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
    12721 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
    12781 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
    12841 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
    12901 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
    12961 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
    13021 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
    13081 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
    13141 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
    13201 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
    13261 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
    13321 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
    13381 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
    13441 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
    13501 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
    13561 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
    13621 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
    13681 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
    13741 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc
    13801 tcccgatccc cta
//
