LOCUS       sequence-259138-       13589 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral all in one expression of CasRx and guideRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13589)
  TITLE     Unpublished
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 13589)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pAK_DR30_EF1a_CasRx_Hygro
FEATURES             Location/Qualifiers
     source          1..13589
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    61..178
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        220..468
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     220..240
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     391..410
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        470..499
                     /label=RfxCas13d DR30
                     /note="5' processed 30-base direct repeat for an RfxCas13d 
                     gRNA (Konermann et al., 2018)"
     promoter        610..1788
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          841..1779
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1736..1756
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             1807..1827
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1837..4734
                     /codon_start=1
                     /product="RNA-targeting Type VI CRISPR protein from 
                     Ruminococcus flavefaciens strain XPD3002 (Konermann et al.,
                     2018)"
                     /label=RfxCas13d
                     /translation="IEKKKSFAKGMGVKSTLVSGSKVYMTTFAEGSDARLEKIVEGDSI
                     RSVNEGEAFSAEMADKNAGYKIGNAKFSHPKGYAVVANNPLYTGPVQQDMLGLKETLEK
                     RYFGESADGNDNICIQVIHNILDIEKILAEYITNAAYAVNNISGLDKDIIGFGKFSTVY
                     TYDEFKDPEHHRAAFNNNDKLINAIKAQYDEFDNFLDNPRLGYFGQAFFSKEGRNYIIN
                     YGNECYDILALLSGLRHWVVHNNEEESRISRTWLYNLDKNLDNEYISTLNYLYDRITNE
                     LTNSFSKNSAANVNYIAETLGINPAEFAEQYFRFSIMKEQKNLGFNITKLREVMLDRKD
                     MSEIRKNHKVFDSIRTKVYTMMDFVIYRYYIEEDAKVAAANKSLPDNEKSLSEKDIFVI
                     NLRGSFNDDQKDALYYDEANRIWRKLENIMHNIKEFRGNKTREYKKKDAPRLPRILPAG
                     RDVSAFSKLMYALTMFLDGKEINDLLTTLINKFDNIQSFLKVMPLIGVNAKFVEEYAFF
                     KDSAKIADELRLIKSFARMGEPIADARRAMYIDAIRILGTNLSYDELKALADTFSLDEN
                     GNKLKKGKHGMRNFIINNVISNKRFHYLIRYGDPAHLHEIAKNEAVVKFVLGRIADIQK
                     KQGQNGKNQIDRYYETCIGKDKGKSVSEKVDALTKIITGMNYDQFDKKRSVIEDTGREN
                     AEREKFKKIISLYLTVIYHILKNIVNINARYVIGFHCVERDAQLYKEKGYDINLKKLEE
                     KGFSSVTKLCAGIDETAPDKRKDVEKEMAERAKESIDSLESANPKLYANYIKYSDEKKA
                     EEFTRQINREKAKTALNAYLRNTKWNVIIREDLLRIDNKTCTLFRNKAVHLEVARYVHA
                     YINDIAEVNSYFQLYHYIMQRIIMNERYEKSSGKVSEYFDAVNDEKKYNDRLLKLLCVP
                     FGYCIPRFKNLSIEALFDRNEAAKFDKEKKKVSGNS"
     CDS             4744..4764
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4774..4800
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             4816..4869
                     /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             4870..5892
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             4870..5892
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     misc_feature    5902..6490
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5955..5975)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6373..6384)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(6493..6509)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6494..6510)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             7015..7195
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(7221..7238)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    7227..7451
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      7497..7925
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7584..7603)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7794..7815
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(7934..7954)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        7939..8268
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      8119..8254
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     8181..8200
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        8316..8363
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             8382..8756
                     /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    8886..9007
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8923..8942)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8977..8996
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(9056..9072)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9056..9072)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(9069..9091)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    9080..9096
                     /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(9104..9110,9111..9128,9129..9134))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    9149..9170
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(9287..9304)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(9458..10046)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(9538..9557)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(10217..11008,11009..11077))
                     /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     10840..10859
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(11078..11182)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(11257..11276)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        11448..11827
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        11829..12027
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     11982..12002
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             12045..12225
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    12272..12397
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    12890..13123
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             13308..13352
                     /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"
ORIGIN
        1 gtgaacggat cggcactgcg tgcgccaatt ctgcagacaa atggcagtat tcatccacaa
       61 ttttaaaaga aaagggggga ttggggggta cagtgcaggg gaaagaatag tagacataat
      121 agcaacagac atacaaacta aagaattaca aaaacaaatt acaaaaattc aaaattttcg
      181 ggtttattac agggacagca gagatccagt ttggttaatg agggcctatt tcccatgatt
      241 ccttcatatt tgcatatacg atacaaggct gttagagaga taattagaat taatttgact
      301 gtaaacacaa agatattagt acaaaatacg tgacgtagaa agtaataatt tcttgggtag
      361 tttgcagttt taaaattatg ttttaaaatg gactatcata tgcttaccgt aacttgaaag
      421 tatttcgatt tcttggcttt atatatcttg tggaaaggac gaaacaccga acccctacca
      481 actggtcggg gtttgaaacg gagacggttg tacgtctctt ttttattaat gcaaagatgg
      541 ataaagtttt aaacagagag gaatctttgc agctaatgga ccttctaggt cttgaaagga
      601 gtgggaattg gctccggtgc ccgtcagtgg gcagagcgca catcgcccac agtccccgag
      661 aagttggggg gaggggtcgg caattgaacc ggtgcctaga gaaggtggcg cggggtaaac
      721 tgggaaagtg atgtcgtgta ctggctccgc ctttttcccg agggtggggg agaaccgtat
      781 ataagtgcag tagtcgccgt gaacgttctt tttcgcaacg ggtttgccgc cagaacacag
      841 gtaagtgccg tgtgtggttc ccgcgggcct ggcctcttta cgggttatgg cccttgcgtg
      901 ccttgaatta cttccacctg gctgcagtac gtgattcttg atcccgagct tcgggttgga
      961 agtgggtggg agagttcgag gccttgcgct taaggagccc cttcgcctcg tgcttgagtt
     1021 gaggcctggc ctgggcgctg gggccgccgc gtgcgaatct ggtggcacct tcgcgcctgt
     1081 ctcgctgctt tcgataagtc tctagccatt taaaattttt gatgacctgc tgcgacgctt
     1141 tttttctggc aagatagtct tgtaaatgcg ggccaagatc tgcacactgg tatttcggtt
     1201 tttggggccg cgggcggcga cggggcccgt gcgtcccagc gcacatgttc ggcgaggcgg
     1261 ggcctgcgag cgcggccacc gagaatcgga cgggggtagt ctcaagctgg ccggcctgct
     1321 ctggtgcctg gcctcgcgcc gccgtgtatc gccccgccct gggcggcaag gctggcccgg
     1381 tcggcaccag ttgcgtgagc ggaaagatgg ccgcttcccg gccctgctgc agggagctca
     1441 aaatggagga cgcggcgctc gggagagcgg gcgggtgagt cacccacaca aaggaaaagg
     1501 gcctttccgt cctcagccgt cgcttcatgt gactccacgg agtaccgggc gccgtccagg
     1561 cacctcgatt agttctcgag cttttggagt acgtcgtctt taggttgggg ggaggggttt
     1621 tatgcgatgg agtttcccca cactgagtgg gtggagactg aagttaggcc agcttggcac
     1681 ttgatgtaat tctccttgga atttgccctt tttgagtttg gatcttggtt cattctcaag
     1741 cctcagacag tggttcaaag tttttttctt ccatttcagg tgtcgtgacg tacggccacc
     1801 atgagcccca agaagaagag aaaggtggag gccagcatcg aaaaaaaaaa gtccttcgcc
     1861 aagggcatgg gcgtgaagtc cacactcgtg tccggctcca aagtgtacat gacaaccttc
     1921 gccgaaggca gcgacgccag gctggaaaag atcgtggagg gcgacagcat caggagcgtg
     1981 aatgagggcg aggccttcag cgctgaaatg gccgataaaa acgccggcta taagatcggc
     2041 aacgccaaat tcagccatcc taagggctac gccgtggtgg ctaacaaccc tctgtataca
     2101 ggacccgtcc agcaggatat gctcggcctg aaggaaactc tggaaaagag gtacttcggc
     2161 gagagcgctg atggcaatga caatatttgt atccaggtga tccataacat cctggacatt
     2221 gaaaaaatcc tcgccgaata cattaccaac gccgcctacg ccgtcaacaa tatctccggc
     2281 ctggataagg acattattgg attcggcaag ttctccacag tgtataccta cgacgaattc
     2341 aaagaccccg agcaccatag ggccgctttc aacaataacg ataagctcat caacgccatc
     2401 aaggcccagt atgacgagtt cgacaacttc ctcgataacc ccagactcgg ctatttcggc
     2461 caggcctttt tcagcaagga gggcagaaat tacatcatca attacggcaa cgaatgctat
     2521 gacattctgg ccctcctgag cggactgagg cactgggtgg tccataacaa cgaagaagag
     2581 tccaggatct ccaggacctg gctctacaac ctcgataaga acctcgacaa cgaatacatc
     2641 tccaccctca actacctcta cgacaggatc accaatgagc tgaccaactc cttctccaag
     2701 aactccgccg ccaacgtgaa ctatattgcc gaaactctgg gaatcaaccc tgccgaattc
     2761 gccgaacaat atttcagatt cagcattatg aaagagcaga aaaacctcgg attcaatatc
     2821 accaagctca gggaagtgat gctggacagg aaggatatgt ccgagatcag gaaaaatcat
     2881 aaggtgttcg actccatcag gaccaaggtc tacaccatga tggactttgt gatttatagg
     2941 tattacatcg aagaggatgc caaggtggct gccgccaata agtccctccc cgataatgag
     3001 aagtccctga gcgagaagga tatctttgtg attaacctga ggggctcctt caacgacgac
     3061 cagaaggatg ccctctacta cgatgaagct aatagaattt ggagaaagct cgaaaatatc
     3121 atgcacaaca tcaaggaatt taggggaaac aagacaagag agtataagaa gaaggacgcc
     3181 cctagactgc ccagaatcct gcccgctggc cgtgatgttt ccgccttcag caaactcatg
     3241 tatgccctga ccatgttcct ggatggcaag gagatcaacg acctcctgac caccctgatt
     3301 aataaattcg ataacatcca gagcttcctg aaggtgatgc ctctcatcgg agtcaacgct
     3361 aagttcgtgg aggaatacgc ctttttcaaa gactccgcca agatcgccga tgagctgagg
     3421 ctgatcaagt ccttcgctag aatgggagaa cctattgccg atgccaggag ggccatgtat
     3481 atcgacgcca tccgtatttt aggaaccaac ctgtcctatg atgagctcaa ggccctcgcc
     3541 gacacctttt ccctggacga gaacggaaac aagctcaaga aaggcaagca cggcatgaga
     3601 aatttcatta ttaataacgt gatcagcaat aaaaggttcc actacctgat cagatacggt
     3661 gatcctgccc acctccatga gatcgccaaa aacgaggccg tggtgaagtt cgtgctcggc
     3721 aggatcgctg acatccagaa aaaacagggc cagaacggca agaaccagat cgacaggtac
     3781 tacgaaactt gtatcggaaa ggataagggc aagagcgtga gcgaaaaggt ggacgctctc
     3841 acaaagatca tcaccggaat gaactacgac caattcgaca agaaaaggag cgtcattgag
     3901 gacaccggca gggaaaacgc cgagagggag aagtttaaaa agatcatcag cctgtacctc
     3961 accgtgatct accacatcct caagaatatt gtcaatatca acgccaggta cgtcatcgga
     4021 ttccattgcg tcgagcgtga tgctcaactg tacaaggaga aaggctacga catcaatctc
     4081 aagaaactgg aagagaaggg attcagctcc gtcaccaagc tctgcgctgg cattgatgaa
     4141 actgcccccg ataagagaaa ggacgtggaa aaggagatgg ctgaaagagc caaggagagc
     4201 attgacagcc tcgagagcgc caaccccaag ctgtatgcca attacatcaa atacagcgac
     4261 gagaagaaag ccgaggagtt caccaggcag attaacaggg agaaggccaa aaccgccctg
     4321 aacgcctacc tgaggaacac caagtggaat gtgatcatca gggaggacct cctgagaatt
     4381 gacaacaaga catgtaccct gttcagaaac aaggccgtcc acctggaagt ggccaggtat
     4441 gtccacgcct atatcaacga cattgccgag gtcaattcct acttccaact gtaccattac
     4501 atcatgcaga gaattatcat gaatgagagg tacgagaaaa gcagcggaaa ggtgtccgag
     4561 tacttcgacg ctgtgaatga cgagaagaag tacaacgata ggctcctgaa actgctgtgt
     4621 gtgcctttcg gctactgtat ccccaggttt aagaacctga gcatcgaggc cctgttcgat
     4681 aggaacgagg ccgccaagtt cgacaaggag aaaaagaagg tgtccggcaa ttccggatcc
     4741 ggacctaaga aaaagaggaa ggtggcggcc gcttacccat acgatgttcc agattacgct
     4801 gctagcggca gtggagaggg cagaggaagt ctgctaacat gcggtgacgt cgaggagaat
     4861 cctggcccaa tgaagcctga actcaccgcg acgtctgtcg agaagtttct gatcgaaaag
     4921 ttcgacagcg tgtccgacct gatgcagctc tcggagggcg aagaatctcg tgctttcagc
     4981 ttcgatgtag gagggcgtgg atatgtcctg cgggtaaata gctgcgccga tggtttctac
     5041 aaagatcgtt atgtttatcg gcactttgca tcggccgcgc tcccgattcc ggaagtgctt
     5101 gacattgggg aattcagcga gagcctgacc tattgcatct cccgccgtgc acagggtgtc
     5161 acgttgcaag acctgcctga aaccgaactg cccgctgttc tgcagccggt cgcggaggcc
     5221 atggatgcga tcgctgcggc cgatcttagc cagacgagcg ggttcggccc attcggaccg
     5281 caaggaatcg gtcaatacac tacatggcgt gatttcatat gcgcgattgc tgatccccat
     5341 gtgtatcact ggcaaactgt gatggacgac accgtcagtg cgtccgtcgc gcaggctctc
     5401 gatgagctga tgctttgggc cgaggactgc cccgaagtcc ggcacctcgt gcacgcggat
     5461 ttcggctcca acaatgtcct gacggacaat ggccgcataa cagcggtcat tgactggagc
     5521 gaggcgatgt tcggggattc ccaatacgag gtcgccaaca tcttcttctg gaggccgtgg
     5581 ttggcttgta tggagcagca gacgcgctac ttcgagcgga ggcatccgga gcttgcagga
     5641 tcgccgcggc tccgggcgta tatgctccgc attggtcttg accaactcta tcagagcttg
     5701 gttgacggca atttcgatga tgcagcttgg gcgcagggtc gatgcgacgc aatcgtccga
     5761 tccggagccg ggactgtcgg gcgtacacaa atcgcccgca gaagcgcggc cgtctggacc
     5821 gatggctgtg tagaagtact cgccgatagt ggaaaccgac gccccagcac tcgtccgagg
     5881 gcaaaggaat aagtctagaa taatcaacct ctggattaca aaatttgtga aagattgact
     5941 ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg
     6001 tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggttg
     6061 ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg
     6121 tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct cctttccggg
     6181 actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg ccttgcccgc
     6241 tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc ggggaaatca
     6301 tcgtcctttc cttggctgct cgcctgtgtt gccacctgga ttctgcgcgg gacgtccttc
     6361 tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct gctgccggct
     6421 ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga gtcggatctc cctttgggcc
     6481 gcctccccgc atcgataccg tcgacctcga gacctagaaa aacatggagc aatcacaagt
     6541 agcaatacag cagctaccaa tgctgattgt gcctggctag aagcacaaga ggaggaggag
     6601 gtgggttttc cagtcacacc tcaggtacct ttaagaccaa tgacttacaa ggcagctgta
     6661 gatcttagcc actttttaaa agaaaagggg ggactggaag ggctaattca ctcccaacga
     6721 agacaagata tccttgatct gtggatctac cacacacaag gctacttccc tgattggcag
     6781 aactacacac cagggccagg gatcagatat ccactgacct ttggatggtg ctacaagcta
     6841 gtaccagttg agcaagagaa ggtagaagaa gccaatgaag gagagaacac ccgcttgtta
     6901 caccctgtga gcctgcatgg gatggatgac ccggagagag aagtattaga gtggaggttt
     6961 gacagccgcc tagcatttca tcacatggcc cgagagctgc atccggactg tactgggtct
     7021 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
     7081 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
     7141 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagggcc
     7201 cgtttaaacc cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg
     7261 cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata
     7321 aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt
     7381 ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt
     7441 gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc
     7501 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     7561 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     7621 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     7681 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     7741 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     7801 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     7861 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     7921 gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     7981 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     8041 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     8101 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     8161 catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta
     8221 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga
     8281 gcttgtatat ccattttcgg atctgatcag cacgtgttga caattaatca tcggcatagt
     8341 atatcggcat agtataatac gacaaggtga ggaactaaac catggccaag ttgaccagtg
     8401 ccgttccggt gctcaccgcg cgcgacgtcg ccggagcggt cgagttctgg accgaccggc
     8461 tcgggttctc ccgggacttc gtggaggacg acttcgccgg tgtggtccgg gacgacgtga
     8521 ccctgttcat cagcgcggtc caggaccagg tggtgccgga caacaccctg gcctgggtgt
     8581 gggtgcgcgg cctggacgag ctgtacgccg agtggtcgga ggtcgtgtcc acgaacttcc
     8641 gggacgcctc cgggccggcc atgaccgaga tcggcgagca gccgtggggg cgggagttcg
     8701 ccctgcgcga cccggccggc aactgcgtgc acttcgtggc cgaggagcag gactgacacg
     8761 tgctacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     8821 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     8881 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     8941 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     9001 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
     9061 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     9121 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     9181 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     9241 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     9301 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     9361 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     9421 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     9481 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     9541 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     9601 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     9661 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     9721 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     9781 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     9841 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     9901 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     9961 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
    10021 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
    10081 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
    10141 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
    10201 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
    10261 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
    10321 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
    10381 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
    10441 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
    10501 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
    10561 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
    10621 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
    10681 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
    10741 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
    10801 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
    10861 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
    10921 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
    10981 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
    11041 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
    11101 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
    11161 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtcgacggat
    11221 cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg atgccgcata
    11281 gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa
    11341 aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc tgcttagggt
    11401 taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac attgattatt
    11461 gactagttat taatagtaat caattacggg gtcattagtt catagcccat atatggagtt
    11521 ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc
    11581 attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg
    11641 tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat
    11701 gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca
    11761 gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat
    11821 taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt ttgactcacg
    11881 gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc accaaaatca
    11941 acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg gcggtaggcg
    12001 tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct ctctggttag
    12061 accagatctg agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat
    12121 aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact
    12181 agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc gcccgaacag
    12241 ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc ggcttgctga
    12301 agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa ttttgactag
    12361 cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg ggagaattag
    12421 atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata aattaaaaca
    12481 tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc tgttagaaac
    12541 atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga caggatcaga
    12601 agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc aaaggataga
    12661 gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca aaagtaagac
    12721 caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg agggacaatt
    12781 ggagaagtga attatataaa tataaagtag taaaaattga accattagga gtagcaccca
    12841 ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt
    12901 tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg acgctgacgg
    12961 tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg ctgagggcta
    13021 ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag ctccaggcaa
    13081 gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct
    13141 ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc
    13201 tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt aacaattaca
    13261 caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag aatgaacaag
    13321 aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata acaaattggc
    13381 tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta agaatagttt
    13441 ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta tcgtttcaga
    13501 cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa gaaggtggag
    13561 agagagacag agacagatcc attcgatta
//
