质粒名称: | pcDNA3.1 myc-NL-GW |
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出品公司: | 爱迪基因 |
目录编号: | 113446 |
存储实验室: | Erich Wanker |
载体骨架基本信息 |
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载体名称: | pcDNA3.1 |
载体抗性: | 氯霉素和氨苄青霉素 |
空载体大小: | 5428 bp |
载体类型: | 哺乳动物表达,荧光素酶;网关穿梭载体 |
筛选标记: | 新霉素(G418) |
高拷贝/低拷贝: | 高拷贝 |
生长条件: | 37 ℃ |
插入基因信息 |
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启动子 | CMV |
克隆方法: | Gateway克隆 |
5' 测序引物 | GAACGGCAACAAAATTATCGAC |
3' 测序引物 | TAGAAGGCACAGTCGAGGCT |
文献引用方法 |
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材料和方法部分: | pcDNA3.1 myc-NL-GW was a gift from Erich Wanker (Addgene plasmid # 113446 ; http://n2t.net/addgene:113446 ; RRID:Addgene_113446) |
参考文献部分: | LuTHy: a double‐readout bioluminescence‐based two‐hybrid technology for quantitative mapping of protein–protein interactions in mammalian cells. Trepte P, Kruse S, Kostova S, Hoffmann S, Buntru A, Tempelmeier A, Secker C, Diez L, Schulz A, Klockmeier K, Zenkner M, Golusik S, Rau K, Schnoegl S, Garner CC, Wanker EE.. Molecular Systems Biology (2018) 14, e8071 10.15252/msb.20178071 |
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载体基本信息
载体图谱质粒图谱和多克隆位点信息
载体简介
载体序列
LOCUS pcDNA3.1 myc-NL-GW 7607 bp ds-DNA circular SYN 04-SEP-2018 DEFINITION NanoLuc Gateway shuttle vector for N-terminal fusions. ACCESSION . VERSION . KEYWORDS pcDNA3.1 myc-NL-GW SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7607) AUTHORS Trepte P, Kruse S, Kostova S, Hoffmann S, Buntru A, Tempelmeier A, Secker C, Diez L, Schulz A, Klockmeier K, Zenkner M, Golusik S, Rau K, Schnoegl S, Garner CC, Wanker EE. TITLE LuTHy: a double‐readout bioluminescence‐based two‐hybrid technology for quantitative mapping of protein–protein interactions in mammalian cells JOURNAL Molecular Systems Biology (2018) 14, e8071 REFERENCE 2 (bases 1 to 7607) AUTHORS . TITLE Direct Submission JOURNAL Exported Sep 4, 2018 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7607 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 103..482 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 483..686 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 637..657 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 731..750 /label=T7 /note="T7 promoter, forward primer" promoter 731..749 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" CDS 775..804 /codon_start=1 /product="Myc (human c-Myc proto-oncogene) epitope tag" /label=Myc /translation="EQKLISEEDL" CDS 805..1314 /codon_start=1 /product="NanoLuc(R) luciferase" /label=Nluc /note="human codon-optimized" /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR LCERILA" protein_bind 1321..1445 /gene="mutant version of attR" /label=attR1 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" promoter 1470..1500 /label=lac UV5 promoter /note="E. coli lac promoter with an ""up"" mutation" CDS 1554..2234 /codon_start=1 /product="chloramphenicol acetyltransferase" /label=CmR /note="confers resistance to chloramphenicol" /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQAGRNLE DPAY" primer_bind complement(1620..1639) /label=CAT-R /note="Chloramphenicol resistance gene, reverse primer" CDS 2554..2859 /codon_start=1 /gene="ccdB" /product="CcdB, a bacterial toxin that poisons DNA gyrase" /label=ccdB /note="Plasmids containing the ccdB gene cannot be propagated in standard E. coli strains." /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK VPRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI" primer_bind 2738..2757 /label=ccdB-fwd /note="ccdB gene, forward primer" protein_bind complement(2900..3024) /gene="mutant version of attR" /label=attR2 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" primer_bind complement(3066..3083) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 3072..3296 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 3342..3770 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3429..3448) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 3639..3660 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(3779..3799) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 3784..4113 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 3964..4099 /label=SV40 ori /note="SV40 origin of replication" primer_bind 4026..4045 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 4180..4974 /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(4234..4253) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 4844..4863 /label=Neo-F /note="Neomycin resistance gene, forward primer" polyA_signal 5148..5269 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(5185..5204) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 5239..5258 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(5318..5334) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(5318..5334) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(5331..5353) /label=M13/pUC Reverse /note="In lacZ gene" 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)." promoter complement(5366..5396) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5411..5432 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(5549..5566) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(5720..6308) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(5800..5819) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(6479..7339) /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 7102..7121 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(7340..7444) /gene="bla" /label=AmpR promoter primer_bind complement(7519..7538) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" ORIGIN 1 aagctacaac aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt agggttaggc 61 gttttgcgct gcttcgcgat gtacgggcca gatatacgcg ttgacattga ttattgacta 121 gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg 181 ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga 241 cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat 301 gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa 361 gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca 421 tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc gctattacca 481 tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac tcacggggat 541 ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg 601 actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt aggcgtgtac 661 ggtgggaggt ctatataagc agagctctct ggctaactag agaacccact gcttactggc 721 ttatcgaaat taatacgact cactataggg agacccaagc tggctagcac catggaacag 781 aaactgatct ctgaagaaga cctggtcttc acactcgaag atttcgttgg ggactggcga 841 cagacagccg gctacaacct ggaccaagtc cttgaacagg gaggtgtgtc cagtttgttt 901 cagaatctcg gggtgtccgt aactccgatc caaaggattg tcctgagcgg tgaaaatggg 961 ctgaagatcg acatccatgt catcatcccg tatgaaggtc tgagcggcga ccaaatgggc 1021 cagatcgaaa aaatttttaa ggtggtgtac cctgtggatg atcatcactt taaggtgatc 1081 ctgcactatg gcacactggt aatcgacggg gttacgccga acatgatcga ctatttcgga 1141 cggccgtatg aaggcatcgc cgtgttcgac ggcaaaaaga tcactgtaac agggaccctg 1201 tggaacggca acaaaattat cgacgagcgc ctgatcaacc ccgacggctc cctgctgttc 1261 cgagtaacca tcaacggagt gaccggctgg cggctgtgcg aacgcattct ggcgaagctt 1321 acaagtttgt acaaaaaagc tgaacgagaa acgtaaaatg atataaatat caatatatta 1381 aattagattt tgcataaaaa acagactaca taatactgta aaacacaaca tatccagtca 1441 ctatggcggc cgcattaggc accccaggct ttacacttta tgcttccggc tcgtataatg 1501 tgtggatttt gagttaggat ccgtcgagat tttcaggagc taaggaagct aaaatggaga 1561 aaaaaatcac tggatatacc accgttgata tatcccaatg gcatcgtaaa gaacattttg 1621 aggcatttca gtcagttgct caatgtacct ataaccagac cgttcagctg gatattacgg 1681 cctttttaaa gaccgtaaag aaaaataagc acaagtttta tccggccttt attcacattc 1741 ttgcccgcct gatgaatgct catccggaat tccgtatggc aatgaaagac ggtgagctgg 1801 tgatatggga tagtgttcac ccttgttaca ccgttttcca tgagcaaact gaaacgtttt 1861 catcgctctg gagtgaatac cacgacgatt tccggcagtt tctacacata tattcgcaag 1921 atgtggcgtg ttacggtgaa aacctggcct atttccctaa agggtttatt gagaatatgt 1981 ttttcgtctc agccaatccc tgggtgagtt tcaccagttt tgatttaaac gtggccaata 2041 tggacaactt cttcgccccc gttttcacca tgggcaaata ttatacgcaa ggcgacaagg 2101 tgctgatgcc gctggcgatt caggttcatc atgccgtttg tgatggcttc catgtcggca 2161 gaatgcttaa tgaattacaa cagtactgcg atgagtggca ggcggggcgt aatctagagg 2221 atccggctta ctaaaagcca gataacagta tgcgtatttg cgcgctgatt tttgcggtat 2281 aagaatatat actgatatgt atacccgaag tatgtcaaaa agaggtatgc tatgaagcag 2341 cgtattacag tgacagttga cagcgacagc tatcagttgc tcaaggcata tatgatgtca 2401 atatctccgg tctggtaagc acaaccatgc agaatgaagc ccgtcgtctg cgtgccgaac 2461 gctggaaagc ggaaaatcag gaagggatgg ctgaggtcgc ccggtttatt gaaatgaacg 2521 gctcttttgc tgacgagaac aggggctggt gaaatgcagt ttaaggttta cacctataaa 2581 agagagagcc gttatcgtct gtttgtggat gtacagagtg atattattga cacgcccggg 2641 cgacggatgg tgatccccct ggccagtgca cgtctgctgt cagataaagt cccccgtgaa 2701 ctttacccgg tggtgcatat cggggatgaa agctggcgca tgatgaccac cgatatggcc 2761 agtgtgccgg tctccgttat cggggaagaa gtggctgatc tcagccaccg cgaaaatgac 2821 atcaaaaacg ccattaacct gatgttctgg ggaatataaa tgtcaggctc ccttatacac 2881 agccagtctg caggtcgacc atagtgactg gatatgttgt gttttacagt attatgtagt 2941 ctgtttttta tgcaaaatct aatttaatat attgatattt atatcatttt acgtttctcg 3001 ttcagctttc ttgtacaaag tggtgtaact cgagtctaga gggcccgttt aaacccgctg 3061 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc 3121 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc 3181 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa 3241 gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc 3301 tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc 3361 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct 3421 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg 3481 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga 3541 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt 3601 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg 3661 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc 3721 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg 3781 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa 3841 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc 3901 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg 3961 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt 4021 ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga 4081 ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt 4141 ttcggatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg 4201 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag 4261 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt 4321 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta 4381 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg 4441 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt 4501 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat 4561 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg 4621 atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca 4681 gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc 4741 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc 4801 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat 4861 attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc 4921 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga 4981 ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt 5041 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga 5101 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg 5161 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt 5221 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta 5281 taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa 5341 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct 5401 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc 5461 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg 5521 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc 5581 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag 5641 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa 5701 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc 5761 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc 5821 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg 5881 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt 5941 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc 6001 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc 6061 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag 6121 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg 6181 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa 6241 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag 6301 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact 6361 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa 6421 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt 6481 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag 6541 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca 6601 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc 6661 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt 6721 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg 6781 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca 6841 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg 6901 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca 6961 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg 7021 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct 7081 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca 7141 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca 7201 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg 7261 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac 7321 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt 7381 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc 7441 cgcgcacatt tccccgaaaa gtgccacctg acgtcgacgg atcgggagat ctcccgatcc 7501 cctatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agtatctgct 7561 ccctgcttgt gtgttggagg tcgctgagta gtgcgcgagc aaaattt //
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