LOCUS       sequence-396590-       12793 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  EnAsCas12a with a deactivated DNAse domain (D908A), N' 5x tandem 
            ALFA-tags and C' VP64 and NLS for CRISPRa.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12793)
  AUTHORS   Griffith AL, Zheng F, McGee AV, Miller NW, Szegletes ZM, Reint G, 
            Gademann F, Nwolah I, Hegde M, Liu YV, Goodale A, Doench JG
  TITLE     Optimization of Cas12a for multiplexed genome-scale transcriptional 
            activation.
  JOURNAL   Cell Genom. 2023 Sep 1;3(9):100387. doi: 10.1016/j.xgen.2023.100387.
            eCollection 2023 Sep 13.
  PUBMED    37719144
REFERENCE   2  (bases 1 to 12793)
  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: "Cell 
            Genom."; date: "2023-09-1"; volume: "3(9):100387. doi"; pages: " 
            10.1016/j.xgen.2023.100387. eCollection 2023 Sep 13"
COMMENT     Sequence Label: pRDA_816
FEATURES             Location/Qualifiers
     source          1..12793
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(47..66)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        238..617
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..817
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     772..792
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             835..1015
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1062..1187
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1680..1913
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2098..2142
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2440..2557
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2640..2851
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      2856..2865
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2901..2921
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2931..2969
                     /codon_start=1
                     /product="rationally designed epitope tag that forms a 
                     stable alpha-helix (Gotzke et al., 2019)"
                     /label=ALFA-tag
                     /note="It may be beneficial to flank the ALFA-tag sequence 
                     with proline residues to preserve the helical structure."
                     /translation="SRLEEELRRRLTE"
     CDS             2994..3032
                     /codon_start=1
                     /product="rationally designed epitope tag that forms a 
                     stable alpha-helix (Gotzke et al., 2019)"
                     /label=ALFA-tag
                     /note="It may be beneficial to flank the ALFA-tag sequence 
                     with proline residues to preserve the helical structure."
                     /translation="SRLEEELRRRLTE"
     CDS             3057..3095
                     /codon_start=1
                     /product="rationally designed epitope tag that forms a 
                     stable alpha-helix (Gotzke et al., 2019)"
                     /label=ALFA-tag
                     /note="It may be beneficial to flank the ALFA-tag sequence 
                     with proline residues to preserve the helical structure."
                     /translation="SRLEEELRRRLTE"
     CDS             3120..3158
                     /codon_start=1
                     /product="rationally designed epitope tag that forms a 
                     stable alpha-helix (Gotzke et al., 2019)"
                     /label=ALFA-tag
                     /note="It may be beneficial to flank the ALFA-tag sequence 
                     with proline residues to preserve the helical structure."
                     /translation="SRLEEELRRRLTE"
     CDS             3183..3221
                     /codon_start=1
                     /product="rationally designed epitope tag that forms a 
                     stable alpha-helix (Gotzke et al., 2019)"
                     /label=ALFA-tag
                     /note="It may be beneficial to flank the ALFA-tag sequence 
                     with proline residues to preserve the helical structure."
                     /translation="SRLEEELRRRLTE"
     CDS             3252..7169
                     /codon_start=1
                     /product="CRISPR-associated protein from Acidaminococcus 
                     sp. BV3L6 (Zetsche et al., 2015)"
                     /label=AsCpf1
                     /translation="TQFEGFTNLYQVSKTLRFELIPQGKTLKHIQEQGFIEEDKARNDH
                     YKELKPIIDRIYKTYADQCLQLVQLDWENLSAAIDSYRKEKTEETRNALIEEQATYRNA
                     IHDYFIGRTDNLTDAINKRHAEIYKGLFKAELFNGKVLKQLGTVTTTEHENALLRSFDK
                     FTTYFSGFYRNRKNVFSAEDISTAIPHRIVQDNFPKFKENCHIFTRLITAVPSLREHFE
                     NVKKAIGIFVSTSIEEVFSFPFYNQLLTQTQIDLYNQLLGGISREAGTEKIKGLNEVLN
                     LAIQKNDETAHIIASLPHRFIPLFKQILSDRNTLSFILEEFKSDEEVIQSFCKYKTLLR
                     NENVLETAEALFNELNSIDLTHIFISHKKLETISSALCDHWDTLRNALYERRISELTGK
                     ITKSAKEKVQRSLKHEDINLQEIISAAGKELSEAFKQKTSEILSHAHAALDQPLPTTLK
                     KQEEKEILKSQLDSLLGLYHLLDWFAVDESNEVDPEFSARLTGIKLEMEPSLSFYNKAR
                     NYATKKPYSVEKFKLNFQMPTLARGWDVNREKNNGAILFVKNGLYYLGIMPKQKGRYKA
                     LSFEPTEKTSEGFDKMYYDYFPDAAKMIPKCSTQLKAVTAHFQTHTTPILLSNNFIEPL
                     EITKEIYDLNNPEKEPKKFQTAYAKKTGDQKGYREALCKWIDFTRDFLSKYTKTTSIDL
                     SSLRPSSQYKDLGEYYAELNPLLYHISFQRIAEKEIMDAVETGKLYLFQIYNKDFAKGH
                     HGKPNLHTLYWTGLFSPENLAKTSIKLNGQAELFYRPKSRMKRMAHRLGEKMLNKKLKD
                     QKTPIPDTLYQELYDYVNHRLSHDLSDEARALLPNVITKEVSHEIIKDRRFTSDKFFFH
                     VPITLNYQAANSPSKFNQRVNAYLKEHPETPIIGIARGERNLIYITVIDSTGKILEQRS
                     LNTIQQFDYQKKLDNREKERVAARQAWSVVGTIKDLKQGYLSQVIHEIVDLMIHYQAVV
                     VLENLNFGFKSKRTGIAEKAVYQQFEKMLIDKLNCLVLKDYPAEKVGGVLNPYQLTDQF
                     TSFAKMGTQSGFLFYVPAPYTSKIDPLTGFVDPFVWKTIKNHESRKHFLEGFDFLHYDV
                     KTGDFILHFKMNRNLSFQRGLPGFMPAWDIVFEKNETQFDAKGTPFIAGKRIVPVIENH
                     RFTGRYRDLYPANELIALLEEKGIVFRDGSNILPKLLENDDSHAIDTMVALIRSVLQMR
                     NSNAATGEDYINSPVRDLNGVCFDSRFQNPEWPMDADANGAYHIALKGQLLLNHLKESK
                     DLKLQNGISNQDWLAYIQELRN"
     regulatory      6889..6898
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             7200..7349
                     /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             7362..7409
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7416..7469
                     /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             7476..7874
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    7885..8473
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7938..7958)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8356..8367)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             8545..8778
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(8804..8821)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    8810..9034
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      9080..9508
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9167..9186)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9377..9398
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(9517..9537)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        9522..9851
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      9702..9837
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     9764..9783
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        9899..9946
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             9965..10339
                     /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    10469..10590
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(10506..10525)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     10560..10579
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(10639..10655)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(10639..10655)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(10652..10674)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    10663..10679
                     /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(10687..10693,10694..10711,10712..10717))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    10732..10753
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(10870..10887)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11041..11629)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(11121..11140)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(11800..12591,12592..12660))
                     /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     12423..12442
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(12661..12765)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
       61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
      121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
      181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
      241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
      301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
      361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
      661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
      721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
      781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct
      841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
      901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
      961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc
     1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc
     1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa
     1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg
     1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata
     1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc
     1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga
     1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc
     1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca
     1501 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg
     1561 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga
     1621 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata
     1681 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg
     1741 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg
     1801 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag
     1861 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt
     1921 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt
     1981 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt
     2041 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag
     2101 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata
     2161 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta
     2221 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta
     2281 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa
     2341 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt
     2401 gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat
     2461 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa
     2521 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag
     2581 agatccagtt tggttaatta aggtaccggg cagaggtctc gcttttttcg agacaacacg
     2641 ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg gcaattgaac
     2701 gggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt actggctccg
     2761 cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg tgaacgttct
     2821 ttttcgcaac gggtttgccg ccagaacaca gaggagccac catggccaaa cggacagccg
     2881 acggaagcga gttcgagtca ccaaagaaga agcggaaagt cggcaccagc agcagactgg
     2941 aggaggagct gagaagaagg ctgaccgagc caccaagtgg agggtctcct gcctccagac
     3001 tcgaggaaga gctcagaagg agactcaccg aagggtcccc aacatctact gaagaatcta
     3061 ggctcgaaga ggaactcagg aggaggctca cagaaggcgg accttcctca ggagcccctt
     3121 cacggctgga agaagaactg cggaggcgcc tcactgagcc atcttccaca gggggaccag
     3181 ccagtcgcct cgaagaagag ctcaggcgga gactgacaga gggagggccg agctctggcg
     3241 cacccggcag cacacagttc gagggcttta ccaacctgta tcaggtgagc aagacactgc
     3301 ggtttgagct gatcccacag ggcaagaccc tgaagcacat ccaggagcag ggcttcatcg
     3361 aggaggacaa ggcccgcaat gatcactaca aggagctgaa gcccatcatc gatcggatct
     3421 acaagaccta tgccgaccag tgcctgcagc tggtgcagct ggattgggag aacctgagcg
     3481 ccgccatcga ctcctataga aaggagaaaa ccgaggagac aaggaacgcc ctgatcgagg
     3541 agcaggccac atatcgcaat gccatccacg actacttcat cggccggaca gacaacctga
     3601 ccgatgccat caataagaga cacgccgaga tctacaaggg cctgttcaag gccgagctgt
     3661 ttaatggcaa ggtgctgaag cagctgggca ccgtgaccac aaccgagcac gagaacgccc
     3721 tgctgcggag cttcgacaag tttacaacct acttctccgg cttttatcgg aacaggaaga
     3781 acgtgttcag cgccgaggat atcagcacag ccatcccaca ccgcatcgtg caggacaact
     3841 tccccaagtt taaggagaat tgtcacatct tcacacgcct gatcaccgcc gtgcccagcc
     3901 tgcgggagca ctttgagaac gtgaagaagg ccatcggcat cttcgtgagc acctccatcg
     3961 aggaggtgtt ttccttccct ttttataacc agctgctgac acagacccag atcgacctgt
     4021 ataaccagct gctgggagga atctctcggg aggcaggcac cgagaagatc aagggcctga
     4081 acgaggtgct gaatctggcc atccagaaga atgatgagac agcccacatc atcgcctccc
     4141 tgccacacag attcatcccc ctgtttaagc agatcctgtc cgataggaac accctgtctt
     4201 tcatcctgga ggagtttaag agcgacgagg aagtgatcca gtccttctgc aagtacaaga
     4261 cactgctgag aaacgagaac gtgctggaga cagccgaggc cctgtttaac gagctgaaca
     4321 gcatcgacct gacacacatc ttcatcagcc acaagaagct ggagacaatc agcagcgccc
     4381 tgtgcgacca ctgggataca ctgaggaatg ccctgtatga gcggagaatc tccgagctga
     4441 caggcaagat caccaagtct gccaaggaga aggtgcagcg cagcctgaag cacgaggata
     4501 tcaacctgca ggagatcatc tctgccgcag gcaaggagct gagcgaggcc ttcaagcaga
     4561 aaaccagcga gatcctgtcc cacgcacacg ccgccctgga tcagccactg cctacaaccc
     4621 tgaagaagca ggaggagaag gagatcctga agtctcagct ggacagcctg ctgggcctgt
     4681 accacctgct ggactggttt gccgtggatg agtccaacga ggtggacccc gagttctctg
     4741 cccggctgac cggcatcaag ctggagatgg agccttctct gagcttctac aacaaggcca
     4801 gaaattatgc caccaagaag ccctactccg tggagaagtt caagctgaac tttcagatgc
     4861 ctacactggc ccgcggctgg gacgtgaatc gggagaagaa caatggcgcc atcctgtttg
     4921 tgaagaacgg cctgtactat ctgggcatca tgccaaagca gaagggcagg tataaggccc
     4981 tgagcttcga gcccacagag aaaaccagcg agggctttga taagatgtac tatgactact
     5041 tccctgatgc cgccaagatg atcccaaagt gcagcaccca gctgaaggcc gtgacagccc
     5101 actttcagac ccacacaacc cccatcctgc tgtccaacaa tttcatcgag cctctggaga
     5161 tcacaaagga gatctacgac ctgaacaatc ctgagaagga gccaaagaag tttcagacag
     5221 cctacgccaa gaaaaccggc gaccagaagg gctacagaga ggccctgtgc aagtggatcg
     5281 acttcacaag ggattttctg tccaagtata ccaagacaac ctctatcgat ctgtctagcc
     5341 tgcggccatc ctctcagtat aaggacctgg gcgagtacta tgccgagctg aatcccctgc
     5401 tgtaccacat cagcttccag agaatcgccg agaaggagat catggatgcc gtggagacag
     5461 gcaagctgta cctgttccag atctataaca aggactttgc caagggccac cacggcaagc
     5521 ctaatctgca cacactgtat tggaccggcc tgttttctcc agagaacctg gccaagacaa
     5581 gcatcaagct gaatggccag gccgagctgt tctaccgccc taagtccagg atgaagagga
     5641 tggcacaccg gctgggagag aagatgctga acaagaagct gaaggatcag aaaaccccaa
     5701 tccccgacac cctgtaccag gagctgtacg actatgtgaa tcacagactg tcccacgacc
     5761 tgtctgatga ggccagggcc ctgctgccca acgtgatcac caaggaggtg tctcacgaga
     5821 tcatcaagga taggcgcttt accagcgaca agttcttttt ccacgtgcct atcacactga
     5881 actatcaggc cgccaattcc ccatctaagt tcaaccagag ggtgaatgcc tacctgaagg
     5941 agcaccccga gacacctatc atcggcatcg cccggggcga gagaaacctg atctatatca
     6001 cagtgatcga ctccaccggc aagatcctgg agcagcggag cctgaacacc atccagcagt
     6061 ttgattacca gaagaagctg gacaacaggg agaaggagag ggtggcagca aggcaggcct
     6121 ggtctgtggt gggcacaatc aaggatctga agcagggcta tctgagccag gtcatccacg
     6181 agatcgtgga cctgatgatc cactaccagg ccgtggtggt gctggagaac ctgaatttcg
     6241 gctttaagag caagaggacc ggcatcgccg agaaggccgt gtaccagcag ttcgagaaga
     6301 tgctgatcga taagctgaat tgcctggtgc tgaaggacta tccagcagag aaagtgggag
     6361 gcgtgctgaa cccataccag ctgacagacc agttcacctc ctttgccaag atgggcaccc
     6421 agtctggctt cctgttttac gtgcctgccc catatacatc taagatcgat cccctgaccg
     6481 gcttcgtgga ccccttcgtg tggaaaacca tcaagaatca cgagagccgc aagcacttcc
     6541 tggagggctt cgactttctg cactacgacg tgaaaaccgg cgacttcatc ctgcacttta
     6601 agatgaacag aaatctgtcc ttccagaggg gcctgcccgg ctttatgcct gcatgggata
     6661 tcgtgttcga gaagaacgag acacagtttg acgccaaggg cacccctttc atcgccggca
     6721 agagaatcgt gccagtgatc gagaatcaca gattcaccgg cagataccgg gacctgtatc
     6781 ctgccaacga gctgatcgcc ctgctggagg agaagggcat cgtgttcagg gatggctcca
     6841 acatcctgcc aaagctgctg gagaatgacg attctcacgc catcgacacc atggtggccc
     6901 tgatccgcag cgtgctgcag atgcggaact ccaatgccgc cacaggcgag gactatatca
     6961 acagccccgt gcgcgatctg aatggcgtgt gcttcgactc ccggtttcag aacccagagt
     7021 ggcccatgga cgccgatgcc aatggcgcct accacatcgc cctgaagggc cagctgctgc
     7081 tgaatcacct gaaggagagc aaggatctga agctgcagaa cggcatctcc aatcaggact
     7141 ggctggccta catccaggag ctgcgcaacg ggtccgggag cggcgggagc ggagggagcg
     7201 acgcattgga cgattttgat ctggatatgc tgggaagtga cgccctcgat gattttgacc
     7261 ttgacatgct tggttcggat gcccttgatg actttgacct cgacatgctc ggcagtgacg
     7321 cccttgatga tttcgacctg gacatgctga gcggaggctc caaaaggccg gcggccacga
     7381 aaaaggccgg ccaggcaaaa aagaaaaagg gaagcgaggg cagaggaagc cttctaacat
     7441 gcggtgacgt ggaggagaat cccggccctt ccgggatggc caagcctttg tctcaagaag
     7501 aatccaccct cattgaaaga gcaacggcta caatcaacag catccccatc tctgaggact
     7561 acagcgtcgc cagcgcagct ctctctagcg acggccgcat cttcactggt gtcaatgtat
     7621 atcattttac tgggggacct tgtgcagaac tcgtggtgct gggcactgct gctgctgcgg
     7681 cagctggcaa cctgacttgt atcgtcgcga tcggaaatga gaacaggggc atcttgagcc
     7741 cctgcggacg gtgccgacag gtgcttctcg atctgcatcc tgggatcaaa gccatagtga
     7801 aggacagtga tggacagccg acggcagttg ggattcgtga attgctgccc tctggttatg
     7861 tgtgggaggg ctaataaatt cgaaaatcaa cctctggatt acaaaatttg tgaaagattg
     7921 actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct
     7981 ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg
     8041 ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact
     8101 gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc
     8161 gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc
     8221 cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa
     8281 tcatcgtcct ttccttggct gctcgcctgt gttgccacct ggattctgcg cgggacgtcc
     8341 ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg
     8401 gctctgcggc ctcttccgcg tcctcgcctt cgccctcaga cgagtcggat ctccctttgg
     8461 gccgcctccc cgcgtcgact ttaagaccaa tgacttacaa ggcagctgta gatcttagcc
     8521 actttttaaa agaaaagggg ggactggaag ggctaattca ctcccaacga agataagatc
     8581 tgctttttgc ttgtactggg tctctctggt tagaccagat ctgagcctgg gagctctctg
     8641 gctaactagg gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag
     8701 tgtgtgcccg tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag
     8761 tgtggaaaat ctctagcagg gcccgtttaa acccgctgat cagcctcgac tgtgccttct
     8821 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     8881 actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt
     8941 cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagataat
     9001 agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg
     9061 ggctctaggg ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     9121 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     9181 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     9241 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
     9301 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     9361 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     9421 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     9481 ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg
     9541 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     9601 caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     9661 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     9721 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     9781 aggccgcctc tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     9841 gcttttgcaa aaagctcccg ggagcttgta tatccatttt cggatctgat cagcacgtgt
     9901 tgacaattaa tcatcggcat agtatatcgg catagtataa tacgacaagg tgaggaacta
     9961 aaccatggcc aagttgacca gtgccgttcc ggtgctcacc gcgcgcgacg tcgccggagc
    10021 ggtcgagttc tggaccgacc ggctcgggtt ctcccgggac ttcgtggagg acgacttcgc
    10081 cggtgtggtc cgggacgacg tgaccctgtt catcagcgcg gtccaggacc aggtggtgcc
    10141 ggacaacacc ctggcctggg tgtgggtgcg cggcctggac gagctgtacg ccgagtggtc
    10201 ggaggtcgtg tccacgaact tccgggacgc ctccgggccg gccatgaccg agatcggcga
    10261 gcagccgtgg gggcgggagt tcgccctgcg cgacccggcc ggcaactgcg tgcacttcgt
    10321 ggccgaggag caggactgac acgtgctacg agatttcgat tccaccgccg ccttctatga
    10381 aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga
    10441 tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa
    10501 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
    10561 tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta
    10621 gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat
    10681 tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
    10741 ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
    10801 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
    10861 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
    10921 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
    10981 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
    11041 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
    11101 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
    11161 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
    11221 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
    11281 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
    11341 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
    11401 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
    11461 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
    11521 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
    11581 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
    11641 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
    11701 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
    11761 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
    11821 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
    11881 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
    11941 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
    12001 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
    12061 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
    12121 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
    12181 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
    12241 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
    12301 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
    12361 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
    12421 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
    12481 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
    12541 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
    12601 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
    12661 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
    12721 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
    12781 agtgccacct gac
//
