LOCUS       sequence-496073-       14860 bp    DNA     circular SYN 02-JAN-2026
DEFINITION  Lentiviral expression of dCas9-KRAB  with H2B-GFP reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14860)
  TITLE     Brock Lab Plasmids 2025
REFERENCE   2  (bases 1 to 14860)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jan 2, 2026 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: lenti-EF1a-dCas9-KRAB-H2B-GFP
FEATURES             Location/Qualifiers
     source          1..14860
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    671..788
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     CDS             complement(1086..1130)
                     /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    1315..1548
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     misc_feature    2041..2166
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             2213..2393
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     promoter        complement(2411..2609)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2436..2456)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2611..2990
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     3162..3181
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        3256..3360
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3361..3429,3430..4221)
                     /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     complement(3579..3598)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4392..4980
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4881..4900
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5134..5151
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5268..5289
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5304..5309,5310..5327,5328..5334)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5342..5358
                     /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)."
     primer_bind     5347..5369
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5366..5382
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5366..5382
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    5431..5552
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5442..5461)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5496..5515
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(5682..6056)
                     /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"
     promoter        complement(6075..6122)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(6170..6499)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6184..6319
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(6238..6257)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     6484..6504
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(6513..6941)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6623..6644)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6835..6854
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    6987..7211
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     7200..7217
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     LTR             7243..7423
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     7928..7944
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     7929..7945
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    7948..8536
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             8054..8065
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     8463..8483
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(join(8562..9275,9276..9278,9279..9281))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(8565..9275,9276..9278,9279..9281))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(8607..8628)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     8956..8975
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     regulatory      9278..9287
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             complement(9300..9677)
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     CDS             complement(9690..9743)
                     /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             complement(9765..9959)
                     /codon_start=1
                     /product="Kruppel-associated box (KRAB) transcriptional 
                     repression domain from the human zinc finger protein ZNF10 
                     (Margolin et al., 1994)"
                     /label=KRAB
                     /translation="RTLVTFKDVFVDFTREEWKLLDTAQQIVYRNVMLENYKNLVSLGY
                     QLTKPDVILRLEKGEEPWLV"
     CDS             complement(9984..10004)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(join(10059..11573,11574..11576,11577..14132,
                     14133..14135,14136..14159))
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the N863A mutation in the HNH 
                     catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKARGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             complement(14160..14207)
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     promoter        complement(join(14223..14860,1..541))
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          join(14232..14860,1..310)
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     complement(14255..14275)
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
ORIGIN
        1 gccagaaaaa aagcgtcgca gcaggtcatc aaaaatttta aatggctaga gacttatcga
       61 aagcagcgag acaggcgcga aggtgccacc agattcgcac gcggcggccc cagcgcccag
      121 gccaggcctc aactcaagca cgaggcgaag gggctcctta agcgcaaggc ctcgaactct
      181 cccacccact tccaacccga agctcgggat caagaatcac gtactgcagc caggtggaag
      241 taattcaagg cacgcaaggg ccataacccg taaagaggcc aggcccgcgg gaaccacaca
      301 cggcacttac ctgtgttctg gcggcaaacc cgttgcgaaa aagaacgttc acggcgacta
      361 ctgcacttat atacggttct cccccaccct cgggaaaaag gcggagccag tacacgacat
      421 cactttccca gtttaccccg cgccaccttc tctaggcacc ggttcaattg ccgacccctc
      481 cccccaactt ctcggggact gtgggcgatg tgcgctctgc ccactgacgg gcaccggagc
      541 caattcccac tcctttcaag acctagaagg tccattagct gcaaagattc ctctctgttt
      601 aaaactttat ccatctttgc agctagctaa ttaaccaaac tggatctctg ctgtccctgt
      661 aataaacccg aaaattttga atttttgtaa tttgtttttg taattcttta gtttgtatgt
      721 ctgttgctat tatgtctact attctttccc ctgcactgta ccccccaatc cccccttttc
      781 ttttaaaatt gtggatgaat actgccattt gtctgcagaa ttggcgcacg cagtgccgat
      841 ccgttcacta atcgaatgga tctgtctctg tctctctctc caccttcttc ttctattcct
      901 tcgggcctgt cgggtcccct cggggttggg aggtgggtct gaaacgataa tggtgaatat
      961 ccctgcctaa ctctattcac tatagaaagt acagcaaaaa ctattcttaa acctaccaag
     1021 cctcctacta tcattatgaa taattttata taccacagcc aatttgttat gttaaaccaa
     1081 ttccacaaac ttgcccattt atctaattcc aataattctt gttcattctt ttcttgctgg
     1141 ttttgcgatt cttcaattaa ggagtgtatt aagcttgtgt aattgttaat ttctctgtcc
     1201 cactccatcc aggtcgtgtg attccaaatc tgttccagag atttattact ccaactagca
     1261 ttccaaggca cagcagtggt gcaaatgagt tttccagagc aaccccaaat ccccaggagc
     1321 tgttgatcct ttaggtatct ttccacagcc aggattcttg cctggagctg cttgatgccc
     1381 cagactgtga gttgcaacag atgctgttgc gcctcaatag ccctcagcaa attgttctgc
     1441 tgctgcacta taccagacaa taattgtctg gcctgtaccg tcagcgtcat tgacgctgcg
     1501 cccatagtgc ttcctgctgc tcccaagaac ccaaggaaca aagctcctat tcccactgct
     1561 cttttttctc tctgcaccac tcttctcttt gccttggtgg gtgctactcc taatggttca
     1621 atttttacta ctttatattt atataattca cttctccaat tgtccctcat atctcctcct
     1681 ccaggtctga agatcagcgg ccgcttgctg tgcggtggtc ttacttttgt tttgctcttc
     1741 ctctatcttg tctaaagctt ccttggtgtc ttttatctct atcctttgat gcacacaata
     1801 gagggttgct actgtattat ataatgatct aagttcttct gatcctgtct gaagggatgg
     1861 ttgtagctgt cccagtattt gtctacagcc ttctgatgtt tctaacaggc caggattaac
     1921 tgcgaatcgt tctagctccc tgcttgccca tactatatgt tttaatttat attttttctt
     1981 tccccctggc cttaaccgaa ttttttccca tcgcgatcta attctccccc gcttaatact
     2041 gacgctctcg cacccatctc tctccttcta gcctccgcta gtcaaaattt ttggcgtact
     2101 caccagtcgc cgcccctcgc ctcttgccgt gcgcgcttca gcaagccgag tcctgcgtcg
     2161 agagagctcc tctggtttcc ctttcgcttt caagtccctg ttcgggcgcc actgctagag
     2221 attttccaca ctgactaaaa gggtctgagg gatctctagt taccagagtc acacaacaga
     2281 cgggcacaca ctacttgaag cactcaaggc aagctttatt gaggcttaag cagtgggttc
     2341 cctagttagc cagagagctc ccaggctcag atctggtcta accagagaga cccagtacag
     2401 gcaaaacgcg ctgcttatat agacctccca ccgtacacgc ctaccgccca tttgcgtcaa
     2461 tggggcggag ttgttacgac attttggaaa gtcccgttga ttttggtgcc aaaacaaact
     2521 cccattgacg tcaatggggt ggagacttgg aaatccccgt gagtcaaacc gctatccacg
     2581 cccattgatg tactgccaaa accgcatcac catggtaata gcgatgacta atacgtagat
     2641 gtactgccaa gtaggaaagt cccataaggt catgtactgg gcataatgcc aggcgggcca
     2701 tttaccgtca ttgacgtcaa tagggggcgt acttggcata tgatacactt gatgtactgc
     2761 caagtgggca gtttaccgta aatactccac ccattgacgt caatggaaag tccctattgg
     2821 cgttactatg ggaacatacg tcattattga cgtcaatggg cgggggtcgt tgggcggtca
     2881 gccaggcggg ccatttaccg taagttatgt aacgcggaac tccatatatg ggctatgaac
     2941 taatgacccc gtaattgatt actattaata actagtcaat aatcaatgtc aacgcgtata
     3001 tctggcccgt acatcgcgaa gcagcgcaaa acgcctaacc ctaagcagat tcttcatgca
     3061 attgtcggtc aagccttgcc ttgttgtagc ttaaattttg ctcgcgcact actcagcgac
     3121 ctccaacaca caagcaggga gcagatactg gcttaactat gcggcatcag agcagattgt
     3181 actgagagtg caccataggg gatcgggaga tctcccgatc cgtcgacgtc aggtggcact
     3241 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     3301 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     3361 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
     3421 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
     3481 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
     3541 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
     3601 cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
     3661 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
     3721 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
     3781 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
     3841 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
     3901 cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
     3961 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
     4021 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct
     4081 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
     4141 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
     4201 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
     4261 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     4321 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     4381 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     4441 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     4501 gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta
     4561 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     4621 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     4681 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     4741 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     4801 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     4861 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     4921 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     4981 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     5041 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
     5101 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
     5161 gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg
     5221 cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtgagttag
     5281 ctcactcatt aggcacccca ggctttacac tttatgcttc cggctcgtat gttgtgtgga
     5341 attgtgagcg gataacaatt tcacacagga aacagctatg accatgatta cgccaagctc
     5401 tagctagagg tcgacggtat acagacatga taagatacat tgatgagttt ggacaaacca
     5461 caactagaat gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat
     5521 ttgtaaccat tataagctgc aataaacaag ttggggtggg cgaagaactc cagcatgaga
     5581 tccccgcgct ggaggatcat ccagccggcg tcccggaaaa cgattccgaa gcccaacctt
     5641 tcatagaagg cggcggtgga atcgaaatct cgtagcacgt gtcagtcctg ctcctcggcc
     5701 acgaagtgca cgcagttgcc ggccgggtcg cgcagggcga actcccgccc ccacggctgc
     5761 tcgccgatct cggtcatggc cggcccggag gcgtcccgga agttcgtgga cacgacctcc
     5821 gaccactcgg cgtacagctc gtccaggccg cgcacccaca cccaggccag ggtgttgtcc
     5881 ggcaccacct ggtcctggac cgcgctgatg aacagggtca cgtcgtcccg gaccacaccg
     5941 gcgaagtcgt cctccacgaa gtcccgggag aacccgagcc ggtcggtcca gaactcgacc
     6001 gctccggcga cgtcgcgcgc ggtgagcacc ggaacggcac tggtcaactt ggccatggtt
     6061 tagttcctca ccttgtcgta ttatactatg ccgatatact atgccgatga ttaattgtca
     6121 acacgtgctg atcagatccg aaaatggata tacaagctcc cgggagcttt ttgcaaaagc
     6181 ctaggcctcc aaaaaagcct cctcactact tctggaatag ctcagaggca gaggcggcct
     6241 cggcctctgc ataaataaaa aaaattagtc agccatgggg cggagaatgg gcggaactgg
     6301 gcggagttag gggcgggatg ggcggagtta ggggcgggac tatggttgct gactaattga
     6361 gatgcatgct ttgcatactt ctgcctgctg gggagcctgg ggactttcca cacctggttg
     6421 ctgactaatt gagatgcatg ctttgcatac ttctgcctgc tggggagcct ggggactttc
     6481 cacaccctaa ctgacacaca ttccacagaa ttaattcgcg ttaaattttt gttaaatcag
     6541 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
     6601 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
     6661 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
     6721 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
     6781 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
     6841 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
     6901 caccacaccc gccgcgctta atgcgccgct acagggcgcg tggggatacc ccctagagcc
     6961 ccagctggtt ctttccgcct cagaagccat agagcccacc gcatccccag catgcctgct
     7021 attgtcttcc caatcctccc ccttgctgtc ctgccccacc ccacccccca gaatagaatg
     7081 acacctactc agacaatgcg atgcaatttc ctcattttat taggaaagga cagtgggagt
     7141 ggcaccttcc agggtcaagg aaggcacggg ggaggggcaa acaacagatg gctggcaact
     7201 agaaggcaca gtcgaggctg atcagcgggt ttaaacgggc cctgctagag attttccaca
     7261 ctgactaaaa gggtctgagg gatctctagt taccagagtc acacaacaga cgggcacaca
     7321 ctacttgaag cactcaaggc aagctttatt gaggcttaag cagtgggttc cctagttagc
     7381 cagagagctc ccaggctcag atctggtcta accagagaga cccagtacag tccggatgca
     7441 gctctcgggc catgtgatga aatgctaggc ggctgtcaaa cctccactct aatacttctc
     7501 tctccgggtc atccatccca tgcaggctca cagggtgtaa caagcgggtg ttctctcctt
     7561 cattggcttc ttctaccttc tcttgctcaa ctggtactag cttgtagcac catccaaagg
     7621 tcagtggata tctgatccct ggccctggtg tgtagttctg ccaatcaggg aagtagcctt
     7681 gtgtgtggta gatccacaga tcaaggatat cttgtcttcg ttgggagtga attagccctt
     7741 ccagtccccc cttttctttt aaaaagtggc taagatctac agctgccttg taagtcattg
     7801 gtcttaaagg tacctgaggt gtgactggaa aacccacctc ctcctcctct tgtgcttcta
     7861 gccaggcaca atcagcattg gtagctgctg tattgctact tgtgattgct ccatgttttt
     7921 ctaggtctcg aggtcgacgg tatcgatgcg gggaggcggc ccaaagggag atccgactcg
     7981 tctgagggcg aaggcgaaga cgcggaagag gccgcagagc cggcagcagg ccgcgggaag
     8041 gaaggtccgc tggattgagg gccgaaggga cgtagcagaa ggacgtcccg cgcagaatcc
     8101 aggtggcaac acaggcgagc agccaaggaa aggacgatga tttccccgac aacaccacgg
     8161 aattgtcagt gcccaacagc cgagcccctg tccagcagcg ggcaaggcag gcggcgatga
     8221 gttccgccgt ggcaataggg agggggaaag cgaaagtccc ggaaaggagc tgacaggtgg
     8281 tggcaatgcc ccaaccagtg ggggttgcgt cagcaaacac agtgcacacc acgccacgtt
     8341 gcctgacaac gggccacaac tcctcataaa gagacagcaa ccaggattta tacaaggagg
     8401 agaaaatgaa agccatacgg gaagcaatag catgatacaa aggcattaaa gcagcgtatc
     8461 cacatagcgt aaaaggagca acatagttaa gaataccagt caatctttca caaattttgt
     8521 aatccagagg ttgattaccg ataagcttga tatcgttcga attacttgta cagctcgtcc
     8581 atgccgagag tgatcccggc ggcggtcacg aactccagca ggaccatgtg atcgcgcttc
     8641 tcgttggggt ctttgctcag ggcggactgg gtgctcaggt agtggttgtc gggcagcagc
     8701 acggggccgt cgccgatggg ggtgttctgc tggtagtggt cggcgagctg cacgctgccg
     8761 tcctcgatgt tgtggcggat cttgaagttc accttgatgc cgttcttctg cttgtcggcc
     8821 atgatataga cgttgtggct gttgtagttg tactccagct tgtgccccag gatgttgccg
     8881 tcctccttga agtcgatgcc cttcagctcg atgcggttca ccagggtgtc gccctcgaac
     8941 ttcacctcgg cgcgggtctt gtagttgccg tcgtccttga agaagatggt gcgctcctgg
     9001 acgtagcctt cgggcatggc ggacttgaag aagtcgtgct gcttcatgtg gtcggggtag
     9061 cggctgaagc actgcacgcc gtaggtcagg gtggtcacga gggtgggcca gggcacgggc
     9121 agcttgccgg tggtgcagat gaacttcagg gtcagcttgc cgtaggtggc atcgccctcg
     9181 ccctcgccgg acacgctgaa cttgtggccg tttacgtcgc cgtccagctc gaccaggatg
     9241 ggcaccaccc cggtgaacag ctcctcgccc ttgctcacca tggtggcgac cggtggatcc
     9301 ttagcgctgg tgtacttggt gatggcctta gtaccctcgg acacggcgtg cttggccaac
     9361 tccccaggca gcagcaggcg cacggccgtc tggatctccc tggaggtgat ggtcgagcgc
     9421 ttgttgtaat gcgccaggcg ggaagcctca cctgcgatgc gctcgaaaat gtcgttcaca
     9481 aacgaattca tgatgcccat ggccttggac gaaatgccgg tgtcagggtg gacctgcttc
     9541 agaaccttgt acacatagat ggaatagctc tccttgcggc tgcgcttgcg cttcttgccg
     9601 cctttcttct gcgccttagt caccgccttc ttggagccct ttttcggggc gggagcagac
     9661 ttcgctggct ctggcatttc gaaggtcatg ggtccgggat tctcttctac gtctccgcat
     9721 gtgagcaggg aacctcttcc ctcccctgaa cctgtacagt taattaccag ccaaggttct
     9781 tccccctttt cgagcctaag gatcacatcc ggtttcgtca actggtatcc gagactgacg
     9841 aggttcttgt aattctcaag catgacattt cgatacacaa tttgttgcgc agtatccagc
     9901 aatttccact cctcacgtgt aaagtccaca aacacgtcct tgaacgtaac cagtgttcta
     9961 gcccgtccgg atccagcggc cgccaccttc ctctttttct taggtccgct acctcctcct
    10021 ccgcttcctc ctcctccgct tccacctcct ccagcgctgt cgcctcccag ctgagacagg
    10081 tcgatccgtg tctcgtacag gccggtgatg ctctggtgga tcagggtggc gtccagcacc
    10141 tctttggtgc tggtgtacct cttccggtcg atggtggtgt caaagtactt gaaggcggca
    10201 ggggctccca gattggtcag ggtaaacagg tggatgatat tctcggcctg ctctctgatg
    10261 ggcttatccc ggtgcttgtt gtaggcggac agcactttgt ccagattagc gtcggccagg
    10321 atcactctct tggagaactc gctgatctgc tcgatgatct cgtccaggta gtgcttgtgc
    10381 tgttccacaa acagctgttt ctgctcatta tcctcggggg agcccttcag cttctcatag
    10441 tggctggcca ggtacaggaa gttcacatat ttggagggca gggccagttc gtttcccttc
    10501 tgcagttcgc cggcagaggc cagcattctc ttccggccgt tttccagctc gaacagggag
    10561 tacttaggca gcttgatgat caggtccttt ttcacttctt tgtagccctt ggcttccaga
    10621 aagtcgatgg gattcttctc gaagctgctt ctttccatga tggtgatccc cagcagctct
    10681 ttcacactct tcagtttctt ggacttgccc ttttccactt tggccaccac cagcacagaa
    10741 taggccacgg tggggctgtc gaagccgccg tacttcttag ggtcccagtc cttctttctg
    10801 gcgatcagct tatcgctgtt cctcttgggc aggatagact ctttgctgaa gccgcctgtc
    10861 tgcacctcgg tctttttcac gatattcact tggggcatgc tcagcacttt ccgcacggtg
    10921 gcaaaatccc ggcccttatc ccacacgatc tccccggttt cgccgtttgt ctcgatcaga
    10981 ggccgcttcc ggatctcgcc gttggccagg gtaatctcgg tcttgaaaaa gttcatgatg
    11041 ttgctgtaga agaagtactt ggcggtagcc ttgccgattt cctgctcgct cttggcgatc
    11101 atcttccgca cgtcgtacac cttgtagtcg ccgtacacga actcgctttc cagcttaggg
    11161 tactttttga tcagggcggt tcccacgacg gcgttcaggt aggcgtcgtg ggcgtggtgg
    11221 tagttgttga tctcgcgcac tttgtaaaac tggaaatcct tccggaaatc ggacaccagc
    11281 ttggacttca gggtgatcac tttcacttcc cggatcagct tgtcattctc gtcgtactta
    11341 gtgttcatcc gggagtccag gatctgtgcc acgtgctttg tgatctgccg ggtttccacc
    11401 agctgtctct tgatgaagcc ggccttatcc agttcgctca ggccgcctct ctcggccttg
    11461 gtcagattgt cgaactttct ctgggtaatc agcttggcgt tcagcagctg ccgccagtag
    11521 ttcttcatct tcttcacgac ctcttcggag ggcacgttgt cgctcttgcc ccgggccttg
    11581 tcgcttctgg tcagcacctt gttgtcgatg gagtcgtcct tcagaaagct ctgaggcacg
    11641 atgtggtcca catcgtagtc ggacagccgg ttgatgtcca gttcctggtc cacgtacata
    11701 tcccgcccat tctgcaggta gtacaggtac agcttctcgt tctgcagctg ggtgttttcc
    11761 acggggtgtt ctttcaggat ctggctgccc agctctttga tgccctcttc gatccgcttc
    11821 attctctcgc ggctgttctt ctgtcccttc tgggtggtct ggttctctct ggccatttcg
    11881 atcacgatgt tctcgggctt gtgccggccc atcactttca cgagctcgtc caccaccttc
    11941 actgtctgca ggatgccctt cttaatggcg gggctgccgg ccagattggc aatgtgctcg
    12001 tgcaggctat cgccctggcc ggacacctgg gctttctgga tgtcctcttt aaaggtcagg
    12061 ctgtcgtcgt ggatcagctg catgaagttt ctgttggcga agccgtcgga cttcaggaaa
    12121 tccaggattg tcttgccgga ctgcttgtcc cggatgccgt tgatcagctt ccggctcagc
    12181 ctgccccagc cggtgtatct ccgccgcttc agctgcttca tcactttgtc gtcgaacagg
    12241 tgggcatagg ttttcagccg ttcctcgatc atctctctgt cctcaaacag tgtcagggtc
    12301 agcacgatat cttccagaat gtcctcgttt tcctcattgt ccaggaagtc cttgtccttg
    12361 ataattttca gcagatcgtg gtatgtgccc agggaggcgt tgaaccgatc ttccacgccg
    12421 gagatttcca cggagtcgaa gcactcgatt ttcttgaagt agtcctcttt cagctgcttc
    12481 acggtcactt tccggttggt cttgaacagc aggtccacga tggccttttt ctgctcgccg
    12541 ctcaggaagg cgggctttct cattccctcg gtcacgtatt tcactttggt cagctcgtta
    12601 tacacggtga agtactcgta cagcaggctg tgcttgggca gcaccttctc gttgggcagg
    12661 ttcttatcga agttggtcat ccgctcgatg aagctctggg cggaagcgcc cttgtccacc
    12721 acttcctcga agttccaggg ggtgatggtt tcctcgctct ttctggtcat ccaggcgaat
    12781 ctgctgtttc ccctggccag agggcccacg tagtagggga tgcggaaggt caggatcttc
    12841 tcgatctttt cccggttgtc cttcaggaat gggtaaaaat cttcctgccg ccgcagaatg
    12901 gcgtgcagct ctcccaggtg gatctggtgg gggatgctgc cgttgtcgaa ggtccgctgc
    12961 ttccgcagca ggtcctctct gttcagcttc acgagcagtt cctcggtgcc gtccatcttt
    13021 tccaggatgg gcttgatgaa cttgtagaac tcttcctggc tggctccgcc gtcaatgtag
    13081 ccggcgtagc cgttcttgct ctggtcgaag aaaatctctt tgtacttctc aggcagctgc
    13141 tgccgcacga gagctttcag cagggtcagg tcctggtggt gctcgtcgta tctcttgatc
    13201 atagaggcgc tcaggggggc cttggtgatc tcggtgttca ctctcaggat gtcgctcagc
    13261 aggatggcgt cggacaggtt cttggcggcc agaaacaggt cggcgtactg gtcgccgatc
    13321 tgggccagca ggttgtccag gtcgtcgtcg taggtgtcct tgctcagctg cagtttggca
    13381 tcctcggcca ggtcgaagtt gctcttgaag ttgggggtca ggcccaggct cagggcaatc
    13441 aggttgccga acaggccatt cttcttctcg ccgggcagct gggcgatcag attttccagc
    13501 cgtctgctct tgctcagtct ggcagacagg atggccttgg cgtccacgcc gctggcgttg
    13561 atggggtttt cctcgaacag ctggttgtag gtctgcacca gctggatgaa cagcttgtcc
    13621 acgtcgctgt tgtcggggtt caggtcgccc tcgatcagga agtggccccg gaacttgatc
    13681 atgtgggcca gggccagata gatcagccgc aggtcggcct tgtcggtgct gtccaccagt
    13741 ttctttctca ggtggtagat ggtggggtac ttctcgtggt aggccacctc gtccacgatg
    13801 ttgccgaaga tggggtgccg ctcgtgcttc ttatcctctt ccaccaggaa ggactcttcc
    13861 agtctgtgga agaagctgtc gtccaccttg gccatctcgt tgctgaagat ctcttgcaga
    13921 tagcagatcc ggttcttccg tctggtgtat cttcttctgg cggttctctt cagccgggtg
    13981 gcctcggctg tttcgccgct gtcgaacagc agggctccga tcaggttctt cttgatgctg
    14041 tgccggtcgg tgttgcccag caccttgaat ttcttgctgg gcaccttgta ctcgtcggtg
    14101 atcacggccc agcccacaga gttggtgccg atggccaggc cgatgctgta cttcttgtcc
    14161 tttttctttt ttgcctggcc ggcctttttc gtggccgccg gccttttcat ggtggccgta
    14221 cgtcacgaca cctgaaatgg aagaaaaaaa ctttgaacca ctgtctgagg cttgagaatg
    14281 aaccaagatc caaactcaaa aagggcaaat tccaaggaga attacatcaa gtgccaagct
    14341 ggcctaactt cagtctccac ccactcagtg tggggaaact ccatcgcata aaacccctcc
    14401 ccccaaccta aagacgacgt actccaaaag ctcgagaact aatcgaggtg cctggacggc
    14461 gcccggtact ccgtggagtc acatgaagcg acggctgagg acggaaaggc ccttttcctt
    14521 tgtgtgggtg actcacccgc ccgctctccc gagcgccgcg tcctccattt tgagctccct
    14581 gcagcagggc cgggaagcgg ccatctttcc gctcacgcaa ctggtgccga ccgggccagc
    14641 cttgccgccc agggcggggc gatacacggc ggcgcgaggc caggcaccag agcaggccgg
    14701 ccagcttgag actacccccg tccgattctc ggtggccgcg ctcgcaggcc ccgcctcgcc
    14761 gaacatgtgc gctgggacgc acgggccccg tcgccgcccg cggccccaaa aaccgaaata
    14821 ccagtgtgca gatcttggcc cgcatttaca agactatctt
//
