质粒名称: | pHyo094 (eMutaT7) |
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出品公司: | Addgene |
目录编号: | 173124 |
存储实验室: | Seokhee Kim |
载体骨架基本信息 |
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载体名称: | pBAD33 |
载体出品公司: | -- |
空载体大小: | 5340 bp |
完整质粒大小: | 8947 bp |
载体修饰: | -- |
载体类型: | 细菌表达 |
筛选标记: | -- |
载体抗性: | 氯霉素 |
生长温度: | 37 ℃ |
宿主菌: | DH5a |
拷贝数: | 低拷贝 |
插入基因信息 |
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插入基因名称: | pmCDA1-T7RNAP |
物种来源: | Synthetic |
插入大小: | 3607 bp |
基因突变: | -- |
GenBank编号 | -- |
启动子: | -- |
标签/融合蛋白: | -- |
克隆方法: | -- |
5’克隆位点: | -- |
3’克隆位点: | -- |
5’测序引物 | pBAD-F (5'-ATGCCATAGCATTTTTATCC-3') |
3’测序引物 | pBAD-R (5'-GATTTAATCTGTATCAGG-3') |
文献引用方法 |
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材料和方法部分: | pHyo094 (eMutaT7) was a gift from Seokhee Kim (Addgene plasmid # 173124 ; http://n2t.net/addgene:173124 ; RRID:Addgene_173124) |
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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 pHyo094 (eMutaT7) 8956 bp ds-DNA circular SYN 14-SEP-2021 DEFINITION Expresses eMutaT7 for mutagenizing genes downstream from a T7 promoter in e.coli. ACCESSION . VERSION . KEYWORDS pHyo094 (eMutaT7) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8956) AUTHORS Park H, Kim S TITLE Gene-specific mutagenesis enables rapid continuous evolution of enzymes in vivo. JOURNAL Nucleic Acids Res. 2021 Apr 6;49(6):e32. doi: 10.1093/nar/gkaa1231. PUBMED 33406230 REFERENCE 2 (bases 1 to 8956) AUTHORS . TITLE Direct Submission JOURNAL Exported Sep 14, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8956 /organism="synthetic DNA construct" /mol_type="other DNA" terminator 43..70 /label=rrnB T2 terminator /note="transcription terminator T2 from the E. coli rrnB gene" promoter 89..180 /gene="bla" /label=AmpR promoter rep_origin 638..1093 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(725..744) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 935..956 /label=F1ori-F /note="F1 origin, forward primer" CDS complement(1649..2308) /codon_start=1 /gene="cat" /product="chloramphenicol acetyltransferase" /label=CmR /note="confers resistance to chloramphenicol" /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA" primer_bind 2223..2242 /label=CAT-R /note="Chloramphenicol resistance gene, reverse primer" promoter complement(2309..2411) /label=cat promoter /note="promoter of the E. coli cat gene encoding chloramphenicol acetyltransferase" primer_bind complement(2803..2820) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(2937..3482) /direction=LEFT /label=p15A ori /note="Plasmids containing the medium-copy-number p15A origin of replication can be propagated in E. coli cells that contain a second plasmid with the ColE1 origin. " primer_bind complement(3017..3036) /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 3558..3577 /label=pBRrevBam /note="pBR322 vectors, tet region, downstream of BamHI, reverse primer" CDS complement(3757..4635) /codon_start=1 /gene="araC" /product="L-arabinose regulatory protein" /label=araC /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE EKVNDVAVKLS" promoter 4662..4946 /gene="araBAD" /label=araBAD promoter /note="promoter of the L-arabinose operon of E. coli; the araC regulatory gene is transcribed in the opposite direction (Guzman et al., 1995)" primer_bind 4869..4888 /label=pBAD Forward /note="For vectors with E. coli araBAD promoter, forward primer" CDS 8358..8612 /codon_start=1 /product="uracil-DNA glycosylase inhibitor from a Bacillus subtilis bacteriophage (Mol et al., 1995)" /label=UGI /translation="MTNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHT AYDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML" primer_bind complement(8651..8668) /label=pBAD Reverse /note="For vectors with E. coli araBAD promoter, reverse primer" terminator 8821..8907 /gene="Escherichia coli rrnB" /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" ORIGIN 1 gggcaggacg cccgccataa actgccaggc atcaaattaa gcagaaggcc atcctgacgg 61 atggcctttt tgcgtttcta caaactcttt tgtttatttt tctaaataca ttcaaatatg 121 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt 181 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct 241 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca 301 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga 361 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta 421 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc 481 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg 541 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt 601 cagaccaagt ttactcatat atactttaga ttgatttacg cgccctgtag cggcgcatta 661 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg 721 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa 781 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc 841 aaaaaacttg atttgggtga tggttcacgt agtgggccat cgccctgata gacggttttt 901 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca 961 acactcaacc ctatctcggg ctattctttt gatttataag ggattttgcc gatttcggcc 1021 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta 1081 acgtttacaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat 1141 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc 1201 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct 1261 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg 1321 cttcagcaga gcgcagatac caaatactgt ccttctagtg tagccgtagt taggccacca 1381 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtcag 1441 gcatttgaga agcacacggt cacactgctt ccggtagtca ataaaccggt aaaccagcaa 1501 tagacataag cggctattta acgaccctgc cctgaaccga cgaccgggtc gaatttgctt 1561 tcgaatttct gccattcatc cgcttattat cacttattca ggcgtagcaa ccaggcgttt 1621 aagggcacca ataactgcct taaaaaaatt acgccccgcc ctgccactca tcgcagtact 1681 gttgtaattc attaagcatt ctgccgacat ggaagccatc acaaacggca tgatgaacct 1741 gaatcgccag cggcatcagc accttgtcgc cttgcgtata atatttgccc atggtgaaaa 1801 cgggggcgaa gaagttgtcc atattggcca cgtttaaatc aaaactggtg aaactcaccc 1861 agggattggc tgagacgaaa aacatattct caataaaccc tttagggaaa taggccaggt 1921 tttcaccgta acacgccaca tcttgcgaat atatgtgtag aaactgccgg aaatcgtcgt 1981 ggtattcact ccagagcgat gaaaacgttt cagtttgctc atggaaaacg gtgtaacaag 2041 ggtgaacact atcccatatc accagctcac cgtctttcat tgccatacgg aattccggat 2101 gagcattcat caggcgggca agaatgtgaa taaaggccgg ataaaacttg tgcttatttt 2161 tctttacggt ctttaaaaag gccgtaatat ccagctgaac ggtctggtta taggtacatt 2221 gagcaactga ctgaaatgcc tcaaaatgtt ctttacgatg ccattgggat atatcaacgg 2281 tggtatatcc agtgattttt ttctccattt tagcttcctt agctcctgaa aatctcgata 2341 actcaaaaaa tacgcccggt agtgatctta tttcattatg gtgaaagttg gaacctctta 2401 cgtgccgatc aacgtctcat tttcgccaaa agttggccca gggcttcccg gtatcaacag 2461 ggacaccagg atttatttat tctgcgaagt gatcttccgt cacaggtatt tattcggcgc 2521 aaagtgcgtc gggtgatgct gccaacttac tgatttagtg tatgatggtg tttttgaggt 2581 gctccagtgg cttctgtttc tatcagctgt ccctcctgtt cagctactga cggggtggtg 2641 cgtaacggca aaagcaccgc cggacatcag cgctagcgga gtgtatactg gcttactatg 2701 ttggcactga tgagggtgtc agtgaagtgc ttcatgtggc aggagaaaaa aggctgcacc 2761 ggtgcgtcag cagaatatgt gatacaggat atattccgct tcctcgctca ctgactcgct 2821 acgctcggtc gttcgactgc ggcgagcgga aatggcttac gaacggggcg gagatttcct 2881 ggaagatgcc aggaagatac ttaacaggga agtgagaggg ccgcggcaaa gccgtttttc 2941 cataggctcc gcccccctga caagcatcac gaaatctgac gctcaaatca gtggtggcga 3001 aacccgacag gactataaag ataccaggcg tttccccctg gcggctccct cgtgcgctct 3061 cctgttcctg cctttcggtt taccggtgtc attccgctgt tatggccgcg tttgtctcat 3121 tccacgcctg acactcagtt ccgggtaggc agttcgctcc aagctggact gtatgcacga 3181 accccccgtt cagtccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc 3241 ggaaagacat gcaaaagcac cactggcagc agccactggt aattgattta gaggagttag 3301 tcttgaagtc atgcgccggt taaggctaaa ctgaaaggac aagttttggt gactgcgctc 3361 ctccaagcca gttacctcgg ttcaaagagt tggtagctca gagaaccttc gaaaaaccgc 3421 cctgcaaggc ggttttttcg ttttcagagc aagagattac gcgcagacca aaacgatctc 3481 aagaagatca tcttattaat cagataaaat atttctaggc tcatgagccc gaagtggcga 3541 gcccgatctt ccccatcggt gatgtcggcg atataggcgc cagcaaccgc acctgtggcg 3601 ccggtgatgc cggccacgat gcgtccggcg tagaggatct gctcatgttt gacagcttat 3661 catcgatgca taatgtgcct gtcaaatgga cgaagcaggg attctgcaaa ccctatgcta 3721 ctccgtcaag ccgtcaattg tctgattcgt taccaattat gacaacttga cggctacatc 3781 attcactttt tcttcacaac cggcacggaa ctcgctcggg ctggccccgg tgcatttttt 3841 aaatacccgc gagaaataga gttgatcgtc aaaaccaaca ttgcgaccga cggtggcgat 3901 aggcatccgg gtggtgctca aaagcagctt cgcctggctg atacgttggt cctcgcgcca 3961 gcttaagacg ctaatcccta actgctggcg gaaaagatgt gacagacgcg acggcgacaa 4021 gcaaacatgc tgtgcgacgc tggcgatatc aaaattgctg tctgccaggt gatcgctgat 4081 gtactgacaa gcctcgcgta cccgattatc catcggtgga tggagcgact cgttaatcgc 4141 ttccatgcgc cgcagtaaca attgctcaag cagatttatc gccagcagct ccgaatagcg 4201 cccttcccct tgcccggcgt taatgatttg cccaaacagg tcgctgaaat gcggctggtg 4261 cgcttcatcc gggcgaaaga accccgtatt ggcaaatatt gacggccagt taagccattc 4321 atgccagtag gcgcgcggac gaaagtaaac ccactggtga taccattcgc gagcctccgg 4381 atgacgaccg tagtgatgaa tctctcctgg cgggaacagc aaaatatcac ccggtcggca 4441 aacaaattct cgtccctgat ttttcaccac cccctgaccg cgaatggtga gattgagaat 4501 ataacctttc attcccagcg gtcggtcgat aaaaaaatcg agataaccgt tggcctcaat 4561 cggcgttaaa cccgccacca gatgggcatt aaacgagtat cccggcagca ggggatcatt 4621 ttgcgcttca gccatacttt tcatactccc gccattcaga gaagaaacca attgtccata 4681 ttgcatcaga cattgccgtc actgcgtctt ttactggctc ttctcgctaa ccaaaccggt 4741 aaccccgctt attaaaagca ttctgtaaca aagcgggacc aaagccatga caaaaacgcg 4801 taacaaaagt gtctataatc acggcagaaa agtccacatt gattatttgc acggcgtcac 4861 actttgctat gccatagcat ttttatccat aagattagcg gatcctacct gacgcttttt 4921 atcgcaactc tctactgttt ctccataccc gtttttttgg gctagcgaat tcgagctcag 4981 gaggaattca ccatggcatt atcatatgat gacagacgcc gagtacgtgc gcattcatga 5041 gaaactggat atttacacct tcaagaagca gttcttcaac aacaagaaat ctgtgtcaca 5101 ccgctgctac gtgctgtttg agttgaagcg aaggggcgaa agaagggctt gcttttgggg 5161 ctatgccgtc aacaagcccc aaagtggcac cgagagagga atacacgctg agatattcag 5221 tatccgaaag gtggaagagt atcttcggga taatcctggg cagtttacga tcaactggta 5281 ttccagctgg agtccttgcg ctgattgtgc cgagaaaatt ctggaatggt ataatcagga 5341 acttcgggga aacgggcaca cattgaaaat ctgggcctgc aagctgtact acgagaagaa 5401 tgcccggaac cagataggac tctggaatct gagggacaat ggtgtaggcc tgaacgtgat 5461 ggtttccgag cactatcagt gttgtcggaa gattttcatc caaagctctc ataaccagct 5521 caatgaaaac cgctggttgg agaaaacact gaaacgtgcg gagaagtgga gatccgagct 5581 gagcatcatg atccaggtca agattctgca taccactaag tctccagccg ttagcgagac 5641 tcccgggacc tcagagtccg ccacacccga aagtaacacg attaacatcg ctaagaacga 5701 cttctctgac atcgaactgg ctgctatccc gttcaacact ctggctgacc attacggtga 5761 gcgtttagct cgcgaacagt tggcccttga gcatgagtct tacgagatgg gtgaagcacg 5821 cttccgcaag atgtttgagc gtcaacttaa agctggtgag gttgcggata acgctgccgc 5881 caagcctctc atcactaccc tactccctaa gatgattgca cgcatcaacg actggtttga 5941 ggaagtgaaa gctaagcgcg gcaagcgccc gacagccttc cagttcctgc aagaaatcaa 6001 gccggaagcc gtagcgtaca tcaccattaa gaccactctg gcttgcctaa ccagtgctga 6061 caatacaacc gttcaggctg tagcaagcgc aatcggtcgg gccattgagg acgaggctcg 6121 cttcggtcgt atccgtgacc ttgaagctaa gcacttcaag aaaaacgttg aggaacaact 6181 caacaagcgc gtagggcacg tctacaagaa agcatttatg caagttgtcg aggctgacat 6241 gctctctaag ggtctactcg gtggcgaggc gtggtcttcg tggcataagg aagactctat 6301 tcatgtagga gtacgctgca tcgagatgct cattgagtca accggaatgg ttagcttaca 6361 ccgccaaaat gctggcgtag taggtcaaga ctctgagact atcgaactcg cacctgaata 6421 cgctgaggct atcgcaaccc gtgcaggtgc gctggctggc atctctccga tgttccaacc 6481 ttgcgtagtt cctcctaagc cgtggactgg cattactggt ggtggctatt gggctaacgg 6541 tcgtcgtcct ctggcgctgg tgcgtactca cagtaagaaa gcactgatgc gctacgaaga 6601 cgtttacatg cctgaggtgt acaaagcgat taacattgcg caaaacaccg catggaaaat 6661 caacaagaaa gtcctagcgg tcgccaacgt aatcaccaag tggaagcatt gtccggtcga 6721 ggacatccct gcgattgagc gtgaagaact cccgatgaaa ccggaagaca tcgacatgaa 6781 tcctgaggct ctcaccgcgt ggaaacgtgc tgccgctgct gtgtaccgca aggacaaggc 6841 tcgcaagtct cgccgtatca gccttgagtt catgcttgag caagccaata agtttgctaa 6901 ccataaggcc atctggttcc cttacaacat ggactggcgc ggtcgtgttt acgctgtgtc 6961 aatgttcaac ccgcaaggta acgatatgac caaaggactg cttacgctgg cgaaaggtaa 7021 accaatcggt aaggaaggtt actactggct gaaaatccac ggtgcaaact gtgcgggtgt 7081 cgataaggtt ccgttccctg agcgcatcaa gttcattgag gaaaaccacg agaacatcat 7141 ggcttgcgct aagtctccac tggagaacac ttggtgggct gagcaagatt ctccgttctg 7201 cttccttgcg ttctgctttg agtacgctgg ggtacagcac cacggcctga gctataactg 7261 ctcccttccg ctggcgtttg acgggtcttg ctctggcatc cagcacttct ccgcgatgct 7321 ccgagatgag gtaggtggtc gcgcggttaa cttgcttcct agtgaaaccg ttcaggacat 7381 ctacgggatt gttgctaaga aagtcaacga gattctacaa gcagacgcaa tcaatgggac 7441 cgataacgaa gtagttaccg tgaccgatga gaacactggt gaaatctctg agaaagtcaa 7501 gctgggcact aaggcactgg ctggtcaatg gctggcttac ggtgttactc gcagtgtgac 7561 taagcgttca gtcatgacgc tggcttacgg gtccaaagag ttcggcttcc gtcaacaagt 7621 gctggaagat accattcagc cagctattga ttccggcaag ggtctgatgt tcactcagcc 7681 gaatcaggct gctggataca tggctaagct gatttgggaa tctgtgagcg tgacggtggt 7741 agctgcggtt gaagcaatga actggcttaa gtctgctgct aagctgctgg ctgctgaggt 7801 caaagataag aagactggag agattcttcg caagcgttgc gctgtgcatt gggtaactcc 7861 tgatggtttc cctgtgtggc aggaatacaa gaagcctatt cagacgcgct tgaacctgat 7921 gttcctcggt cagttccgct tacagcctac cattaacacc aacaaagata gcgagattga 7981 tgcacacaaa caggagtctg gtatcgctcc taactttgta cacagccaag acggtagcca 8041 ccttcgtaag actgtagtgt gggcacacga gaagtacgga atcgaatctt ttgcactgat 8101 tcacgactcc ttcggtacca ttccggctga cgctgcgaac ctgttcaaag cagtgcgcga 8161 aactatggtt gacacctatg agtcttgtga tgtactggct gatttctacg accagttcgc 8221 tgaccagttg cacgagtctc aattggacaa aatgccagca cttccggcta aaggtaactt 8281 gaacctccgt gacatcttag agtcggactt cgcgttcgcg taactgcagc ccggggatcc 8341 tctagagtcg acctgcgatg accaaccttt ccgacatcat agagaaggaa acaggcaaac 8401 agttggtcat ccaagagtcg atactcatgc ttcctgaaga agttgaggag gtcattggga 8461 ataagccgga aagtgacatt ctcgtacaca ctgcgtatga tgagagcacc gatgagaacg 8521 tgatgctgct cacgtcagat gccccagagt acaaaccctg ggctctggtg attcaggact 8581 ctaatggaga gaacaagatc aagatgctgt aaaagcttgg ctgttttggc ggatgagaga 8641 agattttcag cctgatacag attaaatcag aacgcagaag cggtctgata aaacagaatt 8701 tgcctggcgg cagtagcgcg gtggtcccac ctgaccccat gccgaactca gaagtgaaac 8761 gccgtagcgc cgatggtagt gtggggtctc cccatgcgag agtagggaac tgccaggcat 8821 caaataaaac gaaaggctca gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg 8881 gtgaacgctc tcctgagtag gacaaatccg ccgggagcgg atttgaacgt tgcgaagcaa 8941 cggcccggag ggtggc //
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