质粒名称: | pCDF-GFPplus |
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出品公司: | Addgene |
目录编号: | 172718 |
存储实验室: | Tilmann Weber |
载体骨架基本信息 |
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载体名称: | pCDF |
载体出品公司: | Clontech |
空载体大小: | 3725bp |
完整质粒大小: | 4989 bp |
载体类型: | 细菌表达 |
载体抗性: | 氨苄霉素 |
生长温度: | 37 ℃ |
宿主菌: | DH5a |
拷贝数: | 高拷贝 |
插入基因信息 |
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插入基因名称: | GFP plus |
物种来源: | Synthetic |
插入大小: | 717bp |
克隆方法: | Gibson Cloning |
文献引用方法 |
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材料和方法部分: | (Addgene plasmid # 172718 ; http://n2t.net/addgene:172718 ; RRID:Addgene_172718) |
参考文献部分: | A versatile genetic engineering toolkit for E. coli based on CRISPR-prime editing. Tong Y, Jorgensen TS, Whitford CM, Weber T, Lee SY. Nat Commun. 2021 Sep 1;12(1):5206. doi: 10.1038/s41467-021-25541-3. 10.1038/s41467-021-25541-3 PubMed 34471126 |
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pCDF-GFPplus载体质粒基本信息
pCDF-GFPplus质粒图谱载体图谱和pCDF-GFPplus载体序列质粒序列多克隆位点信息
pCDF-GFPplus质粒载体简介
pCDF-GFPplus质粒序列载体序列
LOCUS pCDF-GFPplus 4442 bp DNA circular SYN 25-AUG-2024 DEFINITION Expression of a fast folding GFP variant GFP plus. ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4442) AUTHORS Tong Y, Jorgensen TS, Whitford CM, Weber T, Lee SY TITLE A versatile genetic engineering toolkit for E. coli based on CRISPR-prime editing. JOURNAL Nat Commun. 2021 Sep 1;12(1):5206. doi: 10.1038/s41467-021-25541-3. PUBMED 34471126 REFERENCE 2 (bases 1 to 4442) AUTHORS . TITLE Direct Submission JOURNAL Exported Aug 25, 2024 from SnapGene Server 7.0.3 https://www.biofeng.com COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Nat Commun."; date: "2021-09-1"; volume: "12(1):5206. doi"; pages: " 10.1038/s41467-021-25541-3" FEATURES Location/Qualifiers source 1..4442 /mol_type="other DNA" /organism="synthetic DNA construct" promoter 25..59 /label=BBa_J23106 /note="Anderson constitutive promoter (https://parts.igem.org/Promoters/Catalog/Anderson). Measured strength relative to BBa_J223100: 0.47" RBS 68..79 /note="strong bacterial ribosome binding site (Elowitz and Leibler, 2000)" CDS 86..799 /codon_start=1 /product="GFP variant that folds robustly even when fused to poorly folded proteins (Nager et al., 2011)" /label=superfolder GFP /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE FVTAAGITHGMDELYK" primer_bind complement(116..144) /label=GFP-R /note="GFP, reverse primer. Does NOT anneal to EGFP" primer_bind 739..759 /label=GFP-F /note="GFP, forward primer. Does NOT anneal to EGFP" terminator 872..966 /label=lambda t0 terminator /note="transcription terminator from phage lambda" primer_bind complement(1040..1058) /label=T7 Term /note="T7 terminator, reverse primer" terminator 1054..1101 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" CDS complement(1272..2063) /codon_start=1 /gene="aadA" /product="aminoglycoside adenylyltransferase (Murphy, 1985)" /label=SmR /note="confers resistance to spectinomycin and streptomycin" /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK" promoter complement(2064..2155) /gene="bla" /label=AmpR promoter rep_origin complement(2203..2941) /direction=LEFT /label=CloDF13 ori /note="Plasmids containing the CloDF13 (CDF) origin of replication can be propagated in E. coli cells that contain additional plasmids with compatible origins." CDS complement(3151..4233) /codon_start=1 /gene="lacI" /product="lac repressor" /label=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)." /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR ALADSLMQLARQVSRLESGQ" primer_bind 4195..4214 /label=LacI-R /note="LacI, reverse primer" promoter complement(4234..4311) /gene="lacI" /label=lacI promoter primer_bind 4357..4376 /label=T7 /note="T7 promoter, forward primer" promoter 4357..4375 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" protein_bind 4376..4400 /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)." ORIGIN 1 ggttacggtt gagtaataaa tggatttacg gctagctcag tcctaggtat agtgctagct 61 actagagaaa gaggagaaat actagatgag caaaggagaa gaacttttca ctggagttgt 121 cccaattctt gttgaattag atggtgatgt taatgggcac aaattttctg tcagtggaga 181 gggtgaaggt gatgctacat acggaaaact cacccttaaa tttatttgca ctactggaaa 241 actacctgtt ccatggccaa cacttgtcac tactctgacc tatggtgttc aatgcttttc 301 ccgttatccg gatcacatga aacggcatga ctttttcaag agtgccatgc ccgaaggtta 361 tgtacaggaa cgcactatat ctttcaaaga tgacgggaac tacaagacgc gtgctgaagt 421 caagtttgaa ggtgataccc ttgttaatcg tatcgagtta aagggtattg attttaaaga 481 agatggaaac attctcggac acaaactaga gtacaactat aactcacaca atgtatacat 541 cacggcagac aaacaaaaga atggaatcaa agctaacttc aaaattcgcc acaacattga 601 agatggttcc gttcaactag cagaccatta tcaacaaaat actccaattg gcgatggccc 661 tgtcctttta ccagacaacc attacctgtc gacacaatct gccctttcga aagatcccaa 721 cgaaaagcgt gaccacatgg tccttcttga gtttgtaact gctgctggga ttacacatgg 781 catggatgag ctctacaaat gaagcgcata cctgcaggca tgcaagcttg cggccgcgtc 841 gtgactggga aaaccctggc gactagtctt ggactcctgt tgatagatcc agtaatgacc 901 tcagaactcc atctggattt gttcagaacg ctcggttgcc gccgggcgtt ttttattggt 961 gagaatccag gggtccccaa taattacgat ttaaatttga cataagcctg ttcggttcga 1021 ttaacctagg ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa 1081 cgggtcttga ggggtttttt gctgaaacct caggcatttg agaagcacac ggtcacactg 1141 cttccggtag tcaataaacc ggtaaaccag caatagacat aagcggctat ttaacgaccc 1201 tgccctgaac cgacgaccgg gtcatcgtgg ccggatcttg cggcccctcg gcttgaacga 1261 attgttagac attatttgcc gactaccttg gtgatctcgc ctttcacgta gtggacaaat 1321 tcttccaact gatctgcgcg cgaggccaag cgatcttctt cttgtccaag ataagcctgt 1381 ctagcttcaa gtatgacggg ctgatactgg gccggcaggc gctccattgc ccagtcggca 1441 gcgacatcct tcggcgcgat tttgccggtt actgcgctgt accaaatgcg ggacaacgta 1501 agcactacat ttcgctcatc gccagcccag tcgggcggcg agttccatag cgttaaggtt 1561 tcatttagcg cctcaaatag atcctgttca ggaaccggat caaagagttc ctccgccgct 1621 ggacctacca aggcaacgct atgttctctt gcttttgtca gcaagatagc cagatcaatg 1681 tcgatcgtgg ctggctcgaa gatacctgca agaatgtcat tgcgctgcca ttctccaaat 1741 tgcagttcgc gcttagctgg ataacgccac ggaatgatgt cgtcgtgcac aacaatggtg 1801 acttctacag cgcggagaat ctcgctctct ccaggggaag ccgaagtttc caaaaggtcg 1861 ttgatcaaag ctcgccgcgt tgtttcatca agccttacgg tcaccgtaac cagcaaatca 1921 atatcactgt gtggcttcag gccgccatcc actgcggagc cgtacaaatg tacggccagc 1981 aacgtcggtt cgagatggcg ctcgatgacg ccaactacct ctgatagttg agtcgatact 2041 tcggcgatca ccgcttccct catactcttc ctttttcaat attattgaag catttatcag 2101 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaatagct 2161 agctcactcg gtcgctacgc tccgggcgtg agactgcggc gggcgctgcg gacacataca 2221 aagttaccca cagattccgt ggataagcag gggactaaca tgtgaggcaa aacagcaggg 2281 ccgcgccggt ggcgtttttc cataggctcc gccctcctgc cagagttcac ataaacagac 2341 gcttttccgg tgcatctgtg ggagccgtga ggctcaacca tgaatctgac agtacgggcg 2401 aaacccgaca ggacttaaag atccccaccg tttccggcgg gtcgctccct cttgcgctct 2461 cctgttccga ccctgccgtt taccggatac ctgttccgcc tttctccctt acgggaagtg 2521 tggcgctttc tcatagctca cacactggta tctcggctcg gtgtaggtcg ttcgctccaa 2581 gctgggctgt aagcaagaac tccccgttca gcccgactgc tgcgccttat ccggtaactg 2641 ttcacttgag tccaacccgg aaaagcacgg taaaacgcca ctggcagcag ccattggtaa 2701 ctgggagttc gcagaggatt tgtttagcta aacacgcggt tgctcttgaa gtgtgcgcca 2761 aagtccggct acactggaag gacagatttg gttgctgtgc tctgcgaaag ccagttacca 2821 cggttaagca gttccccaac tgacttaacc ttcgatcaaa ccacctcccc aggtggtttt 2881 ttcgtttaca gggcaaaaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat 2941 cttttctact gaaccgctct agatttcagt gcaatttatc tcttcaaatg tagcacctga 3001 agtcagcccc atacgatata agttgtaatt ctcatgttag tcatgccccg cgcccaccgg 3061 aaggagctga ctgggttgaa ggctctcaag ggcatcggtc gagatcccgg tgcctaatga 3121 gtgagctaac ttacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg 3181 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg 3241 cgccagggtg gtttttcttt tcaccagtga gacgggcaac agctgattgc ccttcaccgc 3301 ctggccctga gagagttgca gcaagcggtc cacgctggtt tgccccagca ggcgaaaatc 3361 ctgtttgatg gtggttaacg gcgggatata acatgagctg tcttcggtat cgtcgtatcc 3421 cactaccgag atgtccgcac caacgcgcag cccggactcg gtaatggcgc gcattgcgcc 3481 cagcgccatc tgatcgttgg caaccagcat cgcagtggga acgatgccct cattcagcat 3541 ttgcatggtt tgttgaaaac cggacatggc actccagtcg ccttcccgtt ccgctatcgg 3601 ctgaatttga ttgcgagtga gatatttatg ccagccagcc agacgcagac gcgccgagac 3661 agaacttaat gggcccgcta acagcgcgat ttgctggtga cccaatgcga ccagatgctc 3721 cacgcccagt cgcgtaccgt cttcatggga gaaaataata ctgttgatgg gtgtctggtc 3781 agagacatca agaaataacg ccggaacatt agtgcaggca gcttccacag caatggcatc 3841 ctggtcatcc agcggatagt taatgatcag cccactgacg cgttgcgcga gaagattgtg 3901 caccgccgct ttacaggctt cgacgccgct tcgttctacc atcgacacca ccacgctggc 3961 acccagttga tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg cgtgcagggc 4021 cagactggag gtggcaacgc caatcagcaa cgactgtttg cccgccagtt gttgtgccac 4081 gcggttggga atgtaattca gctccgccat cgccgcttcc actttttccc gcgttttcgc 4141 agaaacgtgg ctggcctggt tcaccacgcg ggaaacggtc tgataagaga caccggcata 4201 ctctgcgaca tcgtataacg ttactggttt cacattcacc accctgaatt gactctcttc 4261 cgggcgctat catgccatac cgcgaaaggt tttgcgccat tcgatggtgt ccgggatctc 4321 gacgctctcc cttatgcgac tcctgcatta ggaaattaat acgactcact ataggggaat 4381 tgtgagcgga taacaattcc cctgtagaaa taattttgtt taactttaat aaggagatat 4441 ac //
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pCDF-GFPplus质粒载体应用举例
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