LOCUS       sequence-457180-       12672 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expresses inactive hfCas13d in mammalian cells for binding of target
            RNAs in combination with compatible crRNAs. Localized to the 
            nucleus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12672)
  TITLE     Low copy CRISPR-Cas13d mitigates collateral RNA cleavage
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 12672)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: 
            pLentiRNACRISPR_012-TetO-NLS-inactive-hfCas13d-NLS-WPRE-EFS-rtTA3-2A
            -Blast
FEATURES             Location/Qualifiers
     source          1..12672
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..21
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     protein_bind    66..84
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    101..119
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    137..155
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    173..191
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    208..226
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    244..262
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        274..312
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     309..333
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             377..397
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             413..3310
                     /codon_start=1
                     /product="RNA-targeting Type VI CRISPR protein from 
                     Ruminococcus flavefaciens strain XPD3002 (Konermann et al.,
                     2018)"
                     /label=RfxCas13d
                     /translation="IEKKKSFAKGMGVKSTLVSGSKVYMTTFAEGSDARLEKIVEGDSI
                     RSVNEGEAFSAEMADKNAGYKIGNAKFSHPKGYAVVANNPLYTGPVQQDMLGLKETLEK
                     RYFGESADGNDNICIQVIHNILDIEKILVEYITNVVYVVNNISGLDKDIIGFGKFSTVY
                     TYDEFKDPEHHRAAFNNNDKLINAIKAQYDEFDNFLDNPRLGYFGQAFFSKEGRNYIIN
                     YGNECYDILALLSGLAHWVVANNEEESRISRTWLYNLDKNLDNEYISTLNYLYDRITNE
                     LTNSFSKNSAANVNYIAETLGINPAEFAEQYFRFSIMKEQKNLGFNITKLREVMLDRKD
                     MSEIRKNHKVFDSIRTKVYTMMDFVIYRYYIEEDAKVAAANKSLPDNEKSLSEKDIFVI
                     NLRGSFNDDQKDALYYDEANRIWRKLENIMHNIKEFRGNKTREYKKKDAPRLPRILPAG
                     RDVSAFSKLMYALTMFLDGKEINDLLTTLINKFDNIQSFLKVMPLIGVNAKFVEEYAFF
                     KDSAKIADELRLIKSFARMGEPIADARRAMYIDAIRILGTNLSYDELKALADTFSLDEN
                     GNKLKKGKHGMRNFIINNVISNKRFHYLIRYGDPAHLHEIAKNEAVVKFVLGRIADIQK
                     KQGQNGKNQIDRYYETCIGKDKGKSVSEKVDALTKIITGMNYDQFDKKRSVIEDTGREN
                     AEREKFKKIISLYLTVIYHILKNIVNINARYVIGFHCVERDAQLYKEKGYDINLKKLEE
                     KGFSSVTKLCAGIDETAPDKRKDVEKEMAERAKESIDSLESANPKLYANYIKYSDEKKA
                     EEFTRQINREKAKTALNAYLRNTKWNVIIREDLLRIDNKTCTLFANKAVALEVARYVHA
                     YINDIAEVNSYFQLYHYIMQRIIMNERYEKSSGKVSEYFDAVNDEKKYNDRLLKLLCVP
                     FGYCIPRFKNLSIEALFDRNEAAKFDKEKKKVSGNS"
     CDS             3320..3340
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3350..3376
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     misc_feature    3426..4014
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3479..3499)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3897..3908)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4017..4033)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4018..4034)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        4079..4290
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             4315..5019
                     /codon_start=1
                     /product="reverse tetracycline transactivator 3"
                     /label=rtTA3
                     /note="activates expression from Tet-responsive promoters 
                     in the presence of doxycycline"
                     /translation="MSRLDKSKVINGALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHFCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPG"
     CDS             5029..5085
                     /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             5086..5484
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     LTR             5876..6056
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(6082..6099)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6088..6312
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      6358..6786
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6445..6464)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6655..6676
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6795..6815)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6800..7129
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6980..7115
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7042..7061
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        7177..7224
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7243..7617
                     /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    7747..7868
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7784..7803)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7838..7857
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(7917..7933)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7917..7933)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7930..7952)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7941..7957
                     /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(7965..7971,7972..7989,7990..7995))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8010..8031
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(8148..8165)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(8319..8907)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8399..8418)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(9078..9869,9870..9938))
                     /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     9701..9720
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9939..10043)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(10118..10137)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        10309..10688
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        10690..10888
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             10906..11086
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    11133..11258
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    11751..11984
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             12169..12213
                     /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    12511..12628
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 caactttgta tagaaaagtt ggctccgaat tcgcccttca ggtccgaggt tctagacgag
       61 tttactccct atcagtgata gagaacgatg tcgagtttac tccctatcag tgatagagaa
      121 cgtatgtcga gtttactccc tatcagtgat agagaacgta tgtcgagttt actccctatc
      181 agtgatagag aacgtatgtc gagtttatcc ctatcagtga tagagaacgt atgtcgagtt
      241 tactccctat cagtgataga gaacgtatgt cgaggtaggc gtgtacggtg ggaggcctat
      301 ataagcagag ctcgtttagt gaaccgtcag atcgcaaagg gcgaattcga cccaagtttg
      361 tacagccacc atgagcccca agaagaagag aaaggtggag gccagcgaat tcatcgaaaa
      421 aaaaaagtcc ttcgccaagg gcatgggcgt gaagtccaca ctcgtgtccg gctccaaagt
      481 gtacatgaca accttcgccg aaggcagcga cgccaggctg gaaaagatcg tggagggcga
      541 cagcatcagg agcgtgaatg agggcgaggc cttcagcgct gaaatggccg ataaaaacgc
      601 cggctataag atcggcaacg ccaaattcag ccatcctaag ggctacgccg tggtggctaa
      661 caaccctctg tatacaggac ccgtccagca ggatatgctc ggcctgaagg aaactctgga
      721 aaagaggtac ttcggcgaga gcgctgatgg caatgacaat atttgtatcc aggtgatcca
      781 taacatcctg gacattgaga agatcctggt ggagtacatc acaaacgtgg tgtacgtggt
      841 gaacaacatc tccggcctgg ataaggacat tattggattc ggcaagttct ccacagtgta
      901 tacctacgac gaattcaaag accccgagca ccatagggcc gctttcaaca ataacgataa
      961 gctcatcaac gccatcaagg cccagtatga cgagttcgac aacttcctcg ataaccccag
     1021 actcggctat ttcggccagg cctttttcag caaggagggc agaaattaca tcatcaatta
     1081 cggcaacgaa tgctatgaca ttctggccct cctgagcgga ctggcgcact gggtggtcgc
     1141 taacaacgaa gaagagtcca ggatctccag gacctggctc tacaacctcg ataagaacct
     1201 cgacaacgaa tacatctcca ccctcaacta cctctacgac aggatcacca atgagctgac
     1261 caactccttc tccaagaact ccgccgccaa cgtgaactat attgccgaaa ctctgggaat
     1321 caaccctgcc gaattcgccg aacaatattt cagattcagc attatgaaag agcagaaaaa
     1381 cctcggattc aatatcacca agctcaggga agtgatgctg gacaggaagg atatgtccga
     1441 gatcaggaaa aatcataagg tgttcgactc catcaggacc aaggtctaca ccatgatgga
     1501 ctttgtgatt tataggtatt acatcgaaga ggatgccaag gtggctgccg ccaataagtc
     1561 cctccccgat aatgagaagt ccctgagcga gaaggatatc tttgtgatta acctgagggg
     1621 ctccttcaac gacgaccaga aggatgccct ctactacgat gaagctaata gaatttggag
     1681 aaagctcgaa aatatcatgc acaacatcaa ggaatttagg ggaaacaaga caagagagta
     1741 taagaagaag gacgccccta gactgcccag aatcctgccc gctggccgtg atgtttccgc
     1801 cttcagcaaa ctcatgtatg ccctgaccat gttcctggat ggcaaggaga tcaacgacct
     1861 cctgaccacc ctgattaata aattcgataa catccagagc ttcctgaagg tgatgcctct
     1921 catcggagtc aacgctaagt tcgtggagga atacgccttt ttcaaagact ccgccaagat
     1981 cgccgatgag ctgaggctga tcaagtcctt cgctagaatg ggagaaccta ttgccgatgc
     2041 caggagggcc atgtatatcg acgccatccg tattttagga accaacctgt cctatgatga
     2101 gctcaaggcc ctcgccgaca ccttttccct ggacgagaac ggaaacaagc tcaagaaagg
     2161 caagcacggc atgagaaatt tcattattaa taacgtgatc agcaataaaa ggttccacta
     2221 cctgatcaga tacggtgatc ctgcccacct ccatgagatc gccaaaaacg aggccgtggt
     2281 gaagttcgtg ctcggcagga tcgctgacat ccagaaaaaa cagggccaga acggcaagaa
     2341 ccagatcgac aggtactacg aaacttgtat cggaaaggat aagggcaaga gcgtgagcga
     2401 aaaggtggac gctctcacaa agatcatcac cggaatgaac tacgaccaat tcgacaagaa
     2461 aaggagcgtc attgaggaca ccggcaggga aaacgccgag agggagaagt ttaaaaagat
     2521 catcagcctg tacctcaccg tgatctacca catcctcaag aatattgtca atatcaacgc
     2581 caggtacgtc atcggattcc attgcgtcga gcgtgatgct caactgtaca aggagaaagg
     2641 ctacgacatc aatctcaaga aactggaaga gaagggattc agctccgtca ccaagctctg
     2701 cgctggcatt gatgaaactg cccccgataa gagaaaggac gtggaaaagg agatggctga
     2761 aagagccaag gagagcattg acagcctcga gagcgccaac cccaagctgt atgccaatta
     2821 catcaaatac agcgacgaga agaaagccga ggagttcacc aggcagatta acagggagaa
     2881 ggccaaaacc gccctgaacg cctacctgag gaacaccaag tggaatgtga tcatcaggga
     2941 ggacctcctg agaattgaca acaagacatg taccctgttc gcaaacaagg ccgtcgccct
     3001 ggaagtggcc aggtatgtcc acgcctatat caacgacatt gccgaggtca attcctactt
     3061 ccaactgtac cattacatca tgcagagaat tatcatgaat gagaggtacg agaaaagcag
     3121 cggaaaggtg tccgagtact tcgacgctgt gaatgacgag aagaagtaca acgataggct
     3181 cctgaaactg ctgtgtgtgc ctttcggcta ctgtatcccc aggtttaaga acctgagcat
     3241 cgaggccctg ttcgatagga acgaggccgc caagttcgac aaggagaaaa agaaggtgtc
     3301 cggcaattcc ggatccggac ctaagaaaaa gaggaaggtg gcggccgctt acccatacga
     3361 tgttccagat tacgcttaag gtacaaagtg gtgaattccg gcaattcgat atcaagctta
     3421 tcgataatca acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg
     3481 ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt
     3541 cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg
     3601 agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc
     3661 ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc
     3721 tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc
     3781 ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc
     3841 tgctcgcctg tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg
     3901 ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc
     3961 gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcatcgat
     4021 accgtcgacc tcgagtaggt cttgaaagga gtgggaattg gctccggtgc ccgtcagtgg
     4081 gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg caattgatcc
     4141 ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta ctggctccgc
     4201 ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt gaacgttctt
     4261 tttcgcaacg ggtttgccgc cagaacacag gaccggttct agagcgctgc caccatgtct
     4321 aggctggaca agagcaaagt cataaacgga gctctggaat tactcaatgg tgtcggtatc
     4381 gaaggcctga cgacaaggaa actcgctcaa aagctgggag ttgagcagcc taccctgtac
     4441 tggcacgtga agaacaagcg ggccctgctc gatgccctgc caatcgagat gctggacagg
     4501 catcataccc acttctgccc cctggaaggc gagtcatggc aagactttct gcggaacaac
     4561 gccaagtcat accgctgtgc tctcctctca catcgcgacg gggctaaagt gcatctcggc
     4621 acccgcccaa cagagaaaca gtacgaaacc ctggaaaatc agctcgcgtt cctgtgtcag
     4681 caaggcttct ccctggagaa cgcactgtac gctctgtccg ccgtgggcca ctttacactg
     4741 ggctgcgtat tggaggaaca ggagcatcaa gtagcaaaag aggaaagaga gacacctacc
     4801 accgattcta tgcccccact tctgagacaa gcaattgagc tgttcgaccg gcagggagcc
     4861 gaacctgcct tccttttcgg cctggaacta atcatatgtg gcctggagaa acagctaaag
     4921 tgcgaaagcg gcgggccgac cgacgccctt gacgattttg acttagacat gctcccagcc
     4981 gatgcccttg acgattttga ccttgacatg ctccccgggg gatccggcgc aacaaacttc
     5041 tctctgctga aacaagccgg agatgtcgaa gagaatcctg gaccgatggc caagcctttg
     5101 tctcaagaag aatccaccct cattgaaaga gcaacggcta caatcaacag catccccatc
     5161 tctgaagact acagcgtcgc cagcgcagct ctctctagcg acggccgcat cttcactggt
     5221 gtcaatgtat atcattttac tgggggacct tgtgcagaac tcgtggtgct gggcactgct
     5281 gctgctgcgg cagctggcaa cctgacttgt atcgtcgcga tcggaaatga gaacaggggc
     5341 atcttgagcc cctgcggacg gtgccgacag gtgcttctcg atctgcatcc tgggatcaaa
     5401 gccatagtga aggacagtga tggacagccg acggcagttg ggattcgtga attgctgccc
     5461 tctggttatg tgtgggaggg ctaaggtacc tttaagacca atgacttaca aggcagctgt
     5521 agatcttagc cactttttaa aagaaaaggg gggactggaa gggctaattc actcccaacg
     5581 aagacaagat atccttgatc tgtggatcta ccacacacaa ggctacttcc ctgattggca
     5641 gaactacaca ccagggccag ggatcagata tccactgacc tttggatggt gctacaagct
     5701 agtaccagtt gagcaagaga aggtagaaga agccaatgaa ggagagaaca cccgcttgtt
     5761 acaccctgtg agcctgcatg ggatggatga cccggagaga gaagtattag agtggaggtt
     5821 tgacagccgc ctagcatttc atcacatggc ccgagagctg catccggact gtactgggtc
     5881 tctctggtta gaccagatct gagcctggga gctctctggc taactaggga acccactgct
     5941 taagcctcaa taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga
     6001 ctctggtaac tagagatccc tcagaccctt ttagtcagtg tggaaaatct ctagcagggc
     6061 ccgtttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt
     6121 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
     6181 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
     6241 tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg
     6301 tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg
     6361 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     6421 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     6481 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     6541 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     6601 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     6661 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     6721 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     6781 cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     6841 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     6901 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     6961 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     7021 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct
     7081 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg
     7141 agcttgtata tccattttcg gatctgatca gcacgtgttg acaattaatc atcggcatag
     7201 tatatcggca tagtataata cgacaaggtg aggaactaaa ccatggccaa gttgaccagt
     7261 gccgttccgg tgctcaccgc gcgcgacgtc gccggagcgg tcgagttctg gaccgaccgg
     7321 ctcgggttct cccgggactt cgtggaggac gacttcgccg gtgtggtccg ggacgacgtg
     7381 accctgttca tcagcgcggt ccaggaccag gtggtgccgg acaacaccct ggcctgggtg
     7441 tgggtgcgcg gcctggacga gctgtacgcc gagtggtcgg aggtcgtgtc cacgaacttc
     7501 cgggacgcct ccgggccggc catgaccgag atcggcgagc agccgtgggg gcgggagttc
     7561 gccctgcgcg acccggccgg caactgcgtg cacttcgtgg ccgaggagca ggactgacac
     7621 gtgctacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt
     7681 ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc
     7741 caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     7801 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     7861 gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca
     7921 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     7981 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     8041 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
     8101 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     8161 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     8221 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     8281 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     8341 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     8401 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     8461 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     8521 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     8581 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     8641 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     8701 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
     8761 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     8821 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     8881 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     8941 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     9001 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     9061 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     9121 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     9181 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     9241 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     9301 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     9361 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     9421 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     9481 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     9541 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     9601 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     9661 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     9721 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     9781 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     9841 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     9901 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     9961 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
    10021 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga
    10081 tcgggagatc tcccgatccc ctatggtgca ctctcagtac aatctgctct gatgccgcat
    10141 agttaagcca gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca
    10201 aaatttaagc tacaacaagg caaggcttga ccgacaattg catgaagaat ctgcttaggg
    10261 ttaggcgttt tgcgctgctt cgcgatgtac gggccagata tacgcgttga cattgattat
    10321 tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt
    10381 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
    10441 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
    10501 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
    10561 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc
    10621 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
    10681 ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac
    10741 ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc
    10801 aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc
    10861 gtgtacggtg ggaggtctat ataagcagcg cgttttgcct gtactgggtc tctctggtta
    10921 gaccagatct gagcctggga gctctctggc taactaggga acccactgct taagcctcaa
    10981 taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac
    11041 tagagatccc tcagaccctt ttagtcagtg tggaaaatct ctagcagtgg cgcccgaaca
    11101 gggacttgaa agcgaaaggg aaaccagagg agctctctcg acgcaggact cggcttgctg
    11161 aagcgcgcac ggcaagaggc gaggggcggc gactggtgag tacgccaaaa attttgacta
    11221 gcggaggcta gaaggagaga gatgggtgcg agagcgtcag tattaagcgg gggagaatta
    11281 gatcgcgatg ggaaaaaatt cggttaaggc cagggggaaa gaaaaaatat aaattaaaac
    11341 atatagtatg ggcaagcagg gagctagaac gattcgcagt taatcctggc ctgttagaaa
    11401 catcagaagg ctgtagacaa atactgggac agctacaacc atcccttcag acaggatcag
    11461 aagaacttag atcattatat aatacagtag caaccctcta ttgtgtgcat caaaggatag
    11521 agataaaaga caccaaggaa gctttagaca agatagagga agagcaaaac aaaagtaaga
    11581 ccaccgcaca gcaagcggcc gctgatcttc agacctggag gaggagatat gagggacaat
    11641 tggagaagtg aattatataa atataaagta gtaaaaattg aaccattagg agtagcaccc
    11701 accaaggcaa agagaagagt ggtgcagaga gaaaaaagag cagtgggaat aggagctttg
    11761 ttccttgggt tcttgggagc agcaggaagc actatgggcg cagcgtcaat gacgctgacg
    11821 gtacaggcca gacaattatt gtctggtata gtgcagcagc agaacaattt gctgagggct
    11881 attgaggcgc aacagcatct gttgcaactc acagtctggg gcatcaagca gctccaggca
    11941 agaatcctgg ctgtggaaag atacctaaag gatcaacagc tcctggggat ttggggttgc
    12001 tctggaaaac tcatttgcac cactgctgtg ccttggaatg ctagttggag taataaatct
    12061 ctggaacaga tttggaatca cacgacctgg atggagtggg acagagaaat taacaattac
    12121 acaagcttaa tacactcctt aattgaagaa tcgcaaaacc agcaagaaaa gaatgaacaa
    12181 gaattattgg aattagataa atgggcaagt ttgtggaatt ggtttaacat aacaaattgg
    12241 ctgtggtata taaaattatt cataatgata gtaggaggct tggtaggttt aagaatagtt
    12301 tttgctgtac tttctatagt gaatagagtt aggcagggat attcaccatt atcgtttcag
    12361 acccacctcc caaccccgag gggacccgac aggcccgaag gaatagaaga agaaggtgga
    12421 gagagagaca gagacagatc cattcgatta gtgaacggat cggcactgcg tgcgccaatt
    12481 ctgcagacaa atggcagtat tcatccacaa ttttaaaaga aaagggggga ttggggggta
    12541 cagtgcaggg gaaagaatag tagacataat agcaacagac atacaaacta aagaattaca
    12601 aaaacaaatt acaaaaattc aaaattttcg ggtttattac agggacagca gagatccagt
    12661 ttggttaatt aa
//
