| 质粒名称: | LentiSAMPHv2 |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 167934 |
| 存储实验室: | Ronald Germain |
|
载体骨架基本信息 |
|
| 载体名称: | lentiSAMv2 |
| 载体出品公司: | Feng Zhang (Addgene Plasmid #75112) |
| 空载体大小: | 13500bp |
| 完整质粒大小: | 14988bp |
| 载体类型: | 哺乳细胞表达, Lentiviral, CRISPR |
| 筛选标记: | Blasticidin |
| 载体抗性: | 氨苄霉素 |
| 生长温度: | 30 ℃ |
| 宿主菌: | NEB Stable |
| 拷贝数: | 高拷贝 |
|
插入基因信息 |
|
| 启动子: | U6, EF1a |
| 克隆方法: | Gibson Cloning |
|
文献引用方法 |
|
| 材料和方法部分: | LentiSAMPHv2 was a gift from Ronald Germain (Addgene plasmid # 167934 ; http://n2t.net/addgene:167934 ; RRID:Addgene_167934) |
| 参考文献部分: |
Efficient Immune Cell Genome Engineering with Enhanced CRISPR Editing Tools. Chan W, Gottschalk RA, Yao Y, Pomerantz JL, Germain RN. Immunohorizons. 2021 Feb 23;5(2):117-132. doi: 10.4049/immunohorizons.2000082. 10.4049/immunohorizons.2000082 PubMed 33622708
|
买家导航
LentiSAMPHv2载体质粒基本信息
LentiSAMPHv2质粒图谱载体图谱和LentiSAMPHv2载体序列质粒序列多克隆位点信息
LentiSAMPHv2质粒载体简介
LentiSAMPHv2质粒序列载体序列
LOCUS LentiSAMPHv2 14988 bp ds-DNA circular SYN 13-MAY-2021
DEFINITION All-in-one lentiviral CRISPRa gRNA expression with blasticidin
selection marker..
ACCESSION .
VERSION .
KEYWORDS LentiSAMPHv2
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 14988)
AUTHORS Chan W, Gottschalk RA, Yao Y, Pomerantz JL, Germain RN
TITLE Efficient Immune Cell Genome Engineering with Enhanced CRISPR
Editing Tools.
JOURNAL Immunohorizons. 2021 Feb 23;5(2):117-132. doi:
10.4049/immunohorizons.2000082.
PUBMED 33622708
REFERENCE 2 (bases 1 to 14988)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..14988
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind complement(78..97)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 269..648
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 650..848
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 803..823
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
LTR 866..1046
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 1093..1218
/label=HIV-1 Psi
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 1711..1944
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 2129..2173
/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 2471..2588
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
promoter 2638..2878
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
primer_bind 2638..2658
/label=hU6-F
/note="Human U6 promoter, forward primer"
primer_bind 2809..2828
/label=LKO.1 5'
/note="Human U6 promoter, forward primer"
misc_RNA 2924..2942
/label=MS2 stem loop
/note="stem loop that binds the bacteriophage MS2 coat
protein"
misc_RNA 2994..3012
/label=MS2 stem loop
/note="stem loop that binds the bacteriophage MS2 coat
protein"
promoter 3099..3310
/label=EF-1-alpha core promoter
/note="core promoter for human elongation factor
EF-1-alpha"
CDS 3327..3374
/codon_start=1
/product="bipartite nuclear localization signal from
nucleoplasmin"
/label=nucleoplasmin NLS
/translation="KRPAATKKAGQAKKKK"
CDS 3375..7475
/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 7530..7550
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 7575..7724
/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 7746..7802
/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 7803..8189
/codon_start=1
/product="bacteriophage MS2 coat protein"
/label=MS2-N55K
/note="binds with high affinity to a specific stem-loop
structure in the viral RNA (Lim et al., 1994)"
/translation="ASNFTQFVLVDNGGTGDVTVAPSNFANGVAEWISSNSRSQAYKVT
CSVRQSSAQKRKYTIKVEVPKVATQTVGGVELPVAAWRSYLNMELTIPIFATNSDCELI
VKAMQGLLKDGNPIPSAIAANSGIY"
CDS 8244..8264
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 8280..8822
/codon_start=1
/product="C-terminal portion of the p65 subunit of mouse
NF-kappa-B"
/label=p65
/translation="PSGQISNQALALAPSSAPVLAQTMVPSSAMVPLAQPPAPAPVLTP
GPPQSLSAPVPKSTQAGEGTLSEALLHLQFDADEDLGALLGNSTDPGVFTDLASVDNSE
FQQLLNQGVSMSHSTAEPMLMEYPEAITRLVTGSQRPPDPAPTPLGTSGLPNGLSGDED
FSSIADMDFSALLSQISS"
CDS 8847..9218
/codon_start=1
/product="C-terminal activation domain from the human heat
shock transcription factor HSF1"
/label=HSF1
/translation="GFSVDTSALLDLFSPSVTVPDMSLPDLDSSLASIQELLSPQEPPR
PPEAENSSPDSGKQLVHYTAQPLFLLDPGSVDTGSNDLPVLFELGEGSYFSEGDGFAED
PTISLLTGSEPPKAKDPTVS"
CDS 9234..9287
/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 9288..9686
/codon_start=1
/gene="Aspergillus terreus BSD"
/product="blasticidin S deaminase"
/label=BSD
/note="confers resistance to blasticidin"
/translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
KAIVKDSDGQPTAVGIRELLPSGYVWEG"
misc_feature 9711..10299
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
primer_bind complement(9764..9784)
/label=WPRE-R
/note="WPRE, reverse primer"
CDS complement(10182..10193)
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
primer_bind complement(10302..10318)
/label=KS primer
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(10303..10319)
/label=pBluescriptKS
/note="For pBluescript vector"
LTR 10824..11004
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
primer_bind complement(11030..11047)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 11036..11260
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 11306..11734
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(11393..11412)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 11603..11624
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(11743..11763)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 11748..12077
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 11928..12063
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 11990..12009
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
promoter 12125..12172
/label=EM7 promoter
/note="synthetic bacterial promoter "
CDS 12191..12565
/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 12695..12816
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(12732..12751)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 12786..12805
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(12865..12881)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(12865..12881)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(12878..12900)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 12889..12905
/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(12913..12943)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 12958..12979
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(13096..13113)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(13267..13855)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(13347..13366)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(14026..14886)
/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 14649..14668
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(14887..3)
/gene="bla"
/label=AmpR promoter
ORIGIN
1 gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc tcccgatccc
61 ctatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gtatctgctc
121 cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca aaatttaagc tacaacaagg
181 caaggcttga ccgacaattg catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt
241 cgcgatgtac gggccagata tacgcgttga cattgattat tgactagtta ttaatagtaa
301 tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg
361 gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg
421 tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta
481 cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt
541 gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac
601 tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt
661 tggcagtaca tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac
721 cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt
781 cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat
841 ataagcagcg cgttttgcct gtactgggtc tctctggtta gaccagatct gagcctggga
901 gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct
961 tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt
1021 ttagtcagtg tggaaaatct ctagcagtgg cgcccgaaca gggacttgaa agcgaaaggg
1081 aaaccagagg agctctctcg acgcaggact cggcttgctg aagcgcgcac ggcaagaggc
1141 gaggggcggc gactggtgag tacgccaaaa attttgacta gcggaggcta gaaggagaga
1201 gatgggtgcg agagcgtcag tattaagcgg gggagaatta gatcgcgatg ggaaaaaatt
1261 cggttaaggc cagggggaaa gaaaaaatat aaattaaaac atatagtatg ggcaagcagg
1321 gagctagaac gattcgcagt taatcctggc ctgttagaaa catcagaagg ctgtagacaa
1381 atactgggac agctacaacc atcccttcag acaggatcag aagaacttag atcattatat
1441 aatacagtag caaccctcta ttgtgtgcat caaaggatag agataaaaga caccaaggaa
1501 gctttagaca agatagagga agagcaaaac aaaagtaaga ccaccgcaca gcaagcggcc
1561 gctgatcttc agacctggag gaggagatat gagggacaat tggagaagtg aattatataa
1621 atataaagta gtaaaaattg aaccattagg agtagcaccc accaaggcaa agagaagagt
1681 ggtgcagaga gaaaaaagag cagtgggaat aggagctttg ttccttgggt tcttgggagc
1741 agcaggaagc actatgggcg cagcgtcaat gacgctgacg gtacaggcca gacaattatt
1801 gtctggtata gtgcagcagc agaacaattt gctgagggct attgaggcgc aacagcatct
1861 gttgcaactc acagtctggg gcatcaagca gctccaggca agaatcctgg ctgtggaaag
1921 atacctaaag gatcaacagc tcctggggat ttggggttgc tctggaaaac tcatttgcac
1981 cactgctgtg ccttggaatg ctagttggag taataaatct ctggaacaga tttggaatca
2041 cacgacctgg atggagtggg acagagaaat taacaattac acaagcttaa tacactcctt
2101 aattgaagaa tcgcaaaacc agcaagaaaa gaatgaacaa gaattattgg aattagataa
2161 atgggcaagt ttgtggaatt ggtttaacat aacaaattgg ctgtggtata taaaattatt
2221 cataatgata gtaggaggct tggtaggttt aagaatagtt tttgctgtac tttctatagt
2281 gaatagagtt aggcagggat attcaccatt atcgtttcag acccacctcc caaccccgag
2341 gggacccgac aggcccgaag gaatagaaga agaaggtgga gagagagaca gagacagatc
2401 cattcgatta gtgaacggat cggcactgcg tgcgccaatt ctgcagacaa atggcagtat
2461 tcatccacaa ttttaaaaga aaagggggga ttggggggta cagtgcaggg gaaagaatag
2521 tagacataat agcaacagac atacaaacta aagaattaca aaaacaaatt acaaaaattc
2581 aaaattttcg ggtttattac agggacagca gagatccagt ttggttaatt agctagcgag
2641 ggcctatttc ccatgattcc ttcatatttg catatacgat acaaggctgt tagagagata
2701 attggaatta atttgactgt aaacacaaag atattagtac aaaatacgtg acgtagaaag
2761 taataatttc ttgggtagtt tgcagtttta aaattatgtt ttaaaatgga ctatcatatg
2821 cttaccgtaa cttgaaagta tttcgatttc ttggctttat atatcttgtg gaaaggacga
2881 aacaccggag acgggatacc gtctctgttt tagagctagg ccaacatgag gatcacccat
2941 gtctgcaggg cctagcaagt taaaataagg ctagtccgtt atcaacttgg ccaacatgag
3001 gatcacccat gtctgcaggg ccaagtggca ccgagtcggt gctttttttg gatcctaggt
3061 cttgaaagga gtgggaattg gctccggtgc ccgtcagtgg gcagagcgca catcgcccac
3121 agtccccgag aagttggggg gaggggtcgg caattgatcc ggtgcctaga gaaggtggcg
3181 cggggtaaac tgggaaagtg atgtcgtgta ctggctccgc ctttttcccg agggtggggg
3241 agaaccgtat ataagtgcag tagtcgccgt gaacgttctt tttcgcaacg ggtttgccgc
3301 cagaacacag gcgtacggcc accatgaaaa ggccggcggc cacgaaaaag gccggccagg
3361 caaaaaagaa aaaggacaag aagtacagca tcggcctggc catcggcacc aactctgtgg
3421 gctgggccgt gatcaccgac gagtacaagg tgcccagcaa gaaattcaag gtgctgggca
3481 acaccgaccg gcacagcatc aagaagaacc tgatcggagc cctgctgttc gacagcggcg
3541 aaacagccga ggccacccgg ctgaagagaa ccgccagaag aagatacacc agacggaaga
3601 accggatctg ctatctgcaa gagatcttca gcaacgagat ggccaaggtg gacgacagct
3661 tcttccacag actggaagag tccttcctgg tggaagagga taagaagcac gagcggcacc
3721 ccatcttcgg caacatcgtg gacgaggtgg cctaccacga gaagtacccc accatctacc
3781 acctgagaaa gaaactggtg gacagcaccg acaaggccga cctgcggctg atctatctgg
3841 ccctggccca catgatcaag ttccggggcc acttcctgat cgagggcgac ctgaaccccg
3901 acaacagcga cgtggacaag ctgttcatcc agctggtgca gacctacaac cagctgttcg
3961 aggaaaaccc catcaacgcc agcggcgtgg acgccaaggc catcctgtct gccagactga
4021 gcaagagcag acggctggaa aatctgatcg cccagctgcc cggcgagaag aagaatggcc
4081 tgttcggcaa cctgattgcc ctgagcctgg gcctgacccc caacttcaag agcaacttcg
4141 acctggccga ggatgccaaa ctgcagctga gcaaggacac ctacgacgac gacctggaca
4201 acctgctggc ccagatcggc gaccagtacg ccgacctgtt tctggccgcc aagaacctgt
4261 ccgacgccat cctgctgagc gacatcctga gagtgaacac cgagatcacc aaggcccccc
4321 tgagcgcctc tatgatcaag agatacgacg agcaccacca ggacctgacc ctgctgaaag
4381 ctctcgtgcg gcagcagctg cctgagaagt acaaagagat tttcttcgac cagagcaaga
4441 acggctacgc cggctacatt gacggcggag ccagccagga agagttctac aagttcatca
4501 agcccatcct ggaaaagatg gacggcaccg aggaactgct cgtgaagctg aacagagagg
4561 acctgctgcg gaagcagcgg accttcgaca acggcagcat cccccaccag atccacctgg
4621 gagagctgca cgccattctg cggcggcagg aagattttta cccattcctg aaggacaacc
4681 gggaaaagat cgagaagatc ctgaccttcc gcatccccta ctacgtgggc cctctggcca
4741 ggggaaacag cagattcgcc tggatgacca gaaagagcga ggaaaccatc accccctgga
4801 acttcgagga agtggtggac aagggcgctt ccgcccagag cttcatcgag cggatgacca
4861 acttcgataa gaacctgccc aacgagaagg tgctgcccaa gcacagcctg ctgtacgagt
4921 acttcaccgt gtataacgag ctgaccaaag tgaaatacgt gaccgaggga atgagaaagc
4981 ccgccttcct gagcggcgag cagaaaaagg ccatcgtgga cctgctgttc aagaccaacc
5041 ggaaagtgac cgtgaagcag ctgaaagagg actacttcaa gaaaatcgag tgcttcgact
5101 ccgtggaaat ctccggcgtg gaagatcggt tcaacgcctc cctgggcaca taccacgatc
5161 tgctgaaaat tatcaaggac aaggacttcc tggacaatga ggaaaacgag gacattctgg
5221 aagatatcgt gctgaccctg acactgtttg aggacagaga gatgatcgag gaacggctga
5281 aaacctatgc ccacctgttc gacgacaaag tgatgaagca gctgaagcgg cggagataca
5341 ccggctgggg caggctgagc cggaagctga tcaacggcat ccgggacaag cagtccggca
5401 agacaatcct ggatttcctg aagtccgacg gcttcgccaa cagaaacttc atgcagctga
5461 tccacgacga cagcctgacc tttaaagagg acatccagaa agcccaggtg tccggccagg
5521 gcgatagcct gcacgagcac attgccaatc tggccggcag ccccgccatt aagaagggca
5581 tcctgcagac agtgaaggtg gtggacgagc tcgtgaaagt gatgggccgg cacaagcccg
5641 agaacatcgt gatcgaaatg gccagagaga accagaccac ccagaaggga cagaagaaca
5701 gccgcgagag aatgaagcgg atcgaagagg gcatcaaaga gctgggcagc cagatcctga
5761 aagaacaccc cgtggaaaac acccagctgc agaacgagaa gctgtacctg tactacctgc
5821 agaatgggcg ggatatgtac gtggaccagg aactggacat caaccggctg tccgactacg
5881 atgtggacca catcgtgcct cagagctttc tgaaggacga ctccatcgac aacaaggtgc
5941 tgaccagaag cgacaaggcc cggggcaaga gcgacaacgt gccctccgaa gaggtcgtga
6001 agaagatgaa gaactactgg cggcagctgc tgaacgccaa gctgattacc cagagaaagt
6061 tcgacaatct gaccaaggcc gagagaggcg gcctgagcga actggataag gccggcttca
6121 tcaagagaca gctggtggaa acccggcaga tcacaaagca cgtggcacag atcctggact
6181 cccggatgaa cactaagtac gacgagaatg acaagctgat ccgggaagtg aaagtgatca
6241 ccctgaagtc caagctggtg tccgatttcc ggaaggattt ccagttttac aaagtgcgcg
6301 agatcaacaa ctaccaccac gcccacgacg cctacctgaa cgccgtcgtg ggaaccgccc
6361 tgatcaaaaa gtaccctaag ctggaaagcg agttcgtgta cggcgactac aaggtgtacg
6421 acgtgcggaa gatgatcgcc aagagcgagc aggaaatcgg caaggctacc gccaagtact
6481 tcttctacag caacatcatg aactttttca agaccgagat taccctggcc aacggcgaga
6541 tccggaagcg gcctctgatc gagacaaacg gcgaaaccgg ggagatcgtg tgggataagg
6601 gccgggattt tgccaccgtg cggaaagtgc tgagcatgcc ccaagtgaat atcgtgaaaa
6661 agaccgaggt gcagacaggc ggcttcagca aagagtctat cctgcccaag aggaacagcg
6721 ataagctgat cgccagaaag aaggactggg accctaagaa gtacggcggc ttcgacagcc
6781 ccaccgtggc ctattctgtg ctggtggtgg ccaaagtgga aaagggcaag tccaagaaac
6841 tgaagagtgt gaaagagctg ctggggatca ccatcatgga aagaagcagc ttcgagaaga
6901 atcccatcga ctttctggaa gccaagggct acaaagaagt gaaaaaggac ctgatcatca
6961 agctgcctaa gtactccctg ttcgagctgg aaaacggccg gaagagaatg ctggcctctg
7021 ccggcgaact gcagaaggga aacgaactgg ccctgccctc caaatatgtg aacttcctgt
7081 acctggccag ccactatgag aagctgaagg gctcccccga ggataatgag cagaaacagc
7141 tgtttgtgga acagcacaag cactacctgg acgagatcat cgagcagatc agcgagttct
7201 ccaagagagt gatcctggcc gacgctaatc tggacaaagt gctgtccgcc tacaacaagc
7261 accgggataa gcccatcaga gagcaggccg agaatatcat ccacctgttt accctgacca
7321 atctgggagc ccctgccgcc ttcaagtact ttgacaccac catcgaccgg aagaggtaca
7381 ccagcaccaa agaggtgctg gacgccaccc tgatccacca gagcatcacc ggcctgtacg
7441 agacacggat cgacctgtct cagctgggag gcgacagcgc tggaggaggt ggaagcggag
7501 gaggaggaag cggaggagga ggtagcggac ctaagaaaaa gaggaaggtg gcggccgctg
7561 gatccggacg ggctgacgca ttggacgatt ttgatctgga tatgctggga agtgacgccc
7621 tcgatgattt tgaccttgac atgcttggtt cggatgccct tgatgacttt gacctcgaca
7681 tgctcggcag tgacgccctt gatgatttcg acctggacat gctgattaac tgtacagggt
7741 ccggcgcaac aaacttctct ctgctgaaac aagccggaga tgtcgaagag aatcctggac
7801 cggcttcaaa ctttactcag ttcgtgctcg tggacaatgg tgggacaggg gatgtgacag
7861 tggctccttc taatttcgct aatggggtgg cagagtggat cagctccaac tcacggagcc
7921 aggcctacaa ggtgacatgc agcgtcaggc agtctagtgc ccagaagaga aagtatacca
7981 tcaaggtgga ggtccccaaa gtggctaccc agacagtggg cggagtcgaa ctgcctgtcg
8041 ccgcttggag gtcctacctg aacatggagc tcactatccc aattttcgct accaattctg
8101 actgtgaact catcgtgaag gcaatgcagg ggctcctcaa agacggtaat cctatccctt
8161 ccgccatcgc cgctaactca ggtatctaca gcgctggagg aggtggaagc ggaggaggag
8221 gaagcggagg aggaggtagc ggacctaaga aaaagaggaa ggtggcggcc gctggatccc
8281 cttcagggca gatcagcaac caggccctgg ctctggcccc tagctccgct ccagtgctgg
8341 cccagactat ggtgccctct agtgctatgg tgcctctggc ccagccacct gctccagccc
8401 ctgtgctgac cccaggacca ccccagtcac tgagcgctcc agtgcccaag tctacacagg
8461 ccggcgaggg gactctgagt gaagctctgc tgcacctgca gttcgacgct gatgaggacc
8521 tgggagctct gctggggaac agcaccgatc ccggagtgtt cacagatctg gcctccgtgg
8581 acaactctga gtttcagcag ctgctgaatc agggcgtgtc catgtctcat agtacagccg
8641 aaccaatgct gatggagtac cccgaagcca ttacccggct ggtgaccggc agccagcggc
8701 cccccgaccc cgctccaact cccctgggaa ccagcggcct gcctaatggg ctgtccggag
8761 atgaagactt ctcaagcatc gctgatatgg actttagtgc cctgctgtca cagatttcct
8821 ctagtgggca gggaggaggt ggaagcggct tcagcgtgga caccagtgcc ctgctggacc
8881 tgttcagccc ctcggtgacc gtgcccgaca tgagcctgcc tgaccttgac agcagcctgg
8941 ccagtatcca agagctcctg tctccccagg agccccccag gcctcccgag gcagagaaca
9001 gcagcccgga ttcagggaag cagctggtgc actacacagc gcagccgctg ttcctgctgg
9061 accccggctc cgtggacacc gggagcaacg acctgccggt gctgtttgag ctgggagagg
9121 gctcctactt ctccgaaggg gacggcttcg ccgaggaccc caccatctcc ctgctgacag
9181 gctcggagcc tcccaaagcc aaggacccca ctgtctcctg tacaggcagt ggagagggca
9241 gaggaagtct gctaacatgc ggtgacgtcg aggagaatcc tggcccaatg gccaagcctt
9301 tgtctcaaga agaatccacc ctcattgaaa gagcaacggc tacaatcaac agcatcccca
9361 tctctgaaga ctacagcgtc gccagcgcag ctctctctag cgacggccgc atcttcactg
9421 gtgtcaatgt atatcatttt actgggggac cttgtgcaga actcgtggtg ctgggcactg
9481 ctgctgctgc ggcagctggc aacctgactt gtatcgtcgc gatcggaaat gagaacaggg
9541 gcatcttgag cccctgcgga cggtgccgac aggtgcttct cgatctgcat cctgggatca
9601 aagccatagt gaaggacagt gatggacagc cgacggcagt tgggattcgt gaattgctgc
9661 cctctggtta tgtgtgggag ggctaagaat tcgatatcaa gcttatcggt aatcaacctc
9721 tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc
9781 tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca
9841 ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg tggcccgttg
9901 tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact ggttggggca
9961 ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct attgccacgg
10021 cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg ttgggcactg
10081 acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc gcctgtgttg
10141 ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc aatccagcgg
10201 accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc
10261 ctcagacgag tcggatctcc ctttgggccg cctccccgca tcgataccgt cgacctcgag
10321 acctagaaaa acatggagca atcacaagta gcaatacagc agctaccaat gctgattgtg
10381 cctggctaga agcacaagag gaggaggagg tgggttttcc agtcacacct caggtacctt
10441 taagaccaat gacttacaag gcagctgtag atcttagcca ctttttaaaa gaaaaggggg
10501 gactggaagg gctaattcac tcccaacgaa gacaagatat ccttgatctg tggatctacc
10561 acacacaagg ctacttccct gattggcaga actacacacc agggccaggg atcagatatc
10621 cactgacctt tggatggtgc tacaagctag taccagttga gcaagagaag gtagaagaag
10681 ccaatgaagg agagaacacc cgcttgttac accctgtgag cctgcatggg atggatgacc
10741 cggagagaga agtattagag tggaggtttg acagccgcct agcatttcat cacatggccc
10801 gagagctgca tccggactgt actgggtctc tctggttaga ccagatctga gcctgggagc
10861 tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct tgagtgcttc
10921 aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc agaccctttt
10981 agtcagtgtg gaaaatctct agcagggccc gtttaaaccc gctgatcagc ctcgactgtg
11041 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa
11101 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt
11161 aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa
11221 gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc
11281 agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt
11341 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
11401 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
11461 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
11521 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
11581 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
11641 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
11701 aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag
11761 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
11821 agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
11881 tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
11941 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
12001 aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag
12061 gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcagc
12121 acgtgttgac aattaatcat cggcatagta tatcggcata gtataatacg acaaggtgag
12181 gaactaaacc atggccaagt tgaccagtgc cgttccggtg ctcaccgcgc gcgacgtcgc
12241 cggagcggtc gagttctgga ccgaccggct cgggttctcc cgggacttcg tggaggacga
12301 cttcgccggt gtggtccggg acgacgtgac cctgttcatc agcgcggtcc aggaccaggt
12361 ggtgccggac aacaccctgg cctgggtgtg ggtgcgcggc ctggacgagc tgtacgccga
12421 gtggtcggag gtcgtgtcca cgaacttccg ggacgcctcc gggccggcca tgaccgagat
12481 cggcgagcag ccgtgggggc gggagttcgc cctgcgcgac ccggccggca actgcgtgca
12541 cttcgtggcc gaggagcagg actgacacgt gctacgagat ttcgattcca ccgccgcctt
12601 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg
12661 cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg
12721 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
12781 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc
12841 tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
12901 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
12961 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
13021 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
13081 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
13141 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
13201 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
13261 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
13321 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
13381 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
13441 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
13501 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
13561 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
13621 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
13681 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
13741 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
13801 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
13861 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
13921 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
13981 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
14041 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
14101 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
14161 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
14221 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
14281 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
14341 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
14401 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
14461 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
14521 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
14581 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
14641 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
14701 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
14761 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
14821 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
14881 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
14941 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttcc
//
