我们所展示的质粒图谱主要是从文献和开放数据库中收集而来,主要是为了方便研究工作,其中一小部分质粒进行了质量控制,可供科学家使用。
所有的产品都严格仅供科学研究使用,不能应用于临床试验,包括人体摄入、注射或外用的药理学用途。
确保质粒的关键元件正确,但是我们并不能保证实验结果。页面展示的图谱序列为理论序列,可能与测序结果不一致,请自行比对后确定是否满足要求。(如果测过序,本页面一般会提供下载)
开放数据库中的大多数载体序列都没有被完全测序。如果实际序列与参考序列的相似度超过99%,则将其视为正确。
由于科学研究是在探索未知,具有很大的不确定性,在任何情况下,我们都不承担超出质粒本身的额外经济损失或责任。
- 载体名称:
- pPIC9-TSE
- 载体抗性:
- Ampicillin
- 载体长度:
- 10878 bp
- 载体类型:
- Yeast expression vector
- 复制子:
- ori
- 载体来源:
- De Schamphelaire W, Olbrechts A, Meert J, Verhelst K, Roggeman Fonseca M, Vanhoucke M, Beyaert R.
- 启动子:
- AOX1
pPIC9-TSE 载体图谱
质粒操作方法
1. 发货形式:质粒干粉(常温运输,存于-20度,请务必先转化提质粒后使用)
2. 收到质粒干粉后请先5000rpm离心1min,再加入20μl ddH2O溶解质粒;(质粒复测的浓度有时候与标称值差距较大,这可能是因为冻干质粒在管中的位置、复溶效率、测量偏差以及管壁的吸附导致,因此建议先转化提质粒后再使用)
3. 取1支100μl 感受态于冰上解冻10min,加入2μl质粒,再冰浴30min后,42℃热激60s,不要搅动,再冰浴2min;
4. 加入900μl无抗的LB液体培养基,180rpm震荡37℃培养45min (30℃培养1-1.5小时);
5. 6000rpm离心5min,仅留100μl上清液重悬细菌沉淀,并涂布至目标质粒抗性的LB平板上;
6. 将平板倒置37℃培养14h,如果要求是30℃则培养20h; (菌落过多则将质粒稀释后再转化;没有菌落则加入10μl质粒转化;建议不要直接转表达感受态, 要先转克隆感受态,重提质粒后再导入表达感受态);
7. 挑取单菌落至LB液体培养基中,加入对应抗生素,220rpm震荡培养14h,根据实验需要和质粒提取试剂盒说明书提取质粒。
pPIC9-TSE 载体序列
LOCUS 40924_34540 10878 bp DNA circular SYN 18-DEC-2018 DEFINITION Yeast expression vector pPIC9-TSE, complete sequence. ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 10878) AUTHORS De Schamphelaire W, Olbrechts A, Meert J, Verhelst K, Roggeman Fonseca M, Vanhoucke M, Beyaert R. TITLE BCCM/LMBP Plasmid collection JOURNAL Unpublished REFERENCE 2 (bases 1 to 10878) AUTHORS De Schamphelaire W. TITLE Direct Submission JOURNAL Submitted (02-FEB-2017) BCCM/LMBP, Universiteit Gent, Technologiepark 927, 9052, BELGIUM REFERENCE 3 (bases 1 to 10878) TITLE Direct Submission REFERENCE 4 (bases 1 to 10878) AUTHORS . TITLE Direct Submission COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Unpublished" COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted (02-FEB-2017) BCCM/LMBP, Universiteit Gent, Technologiepark 927, 9052, BELGIUM" COMMENT SGRef: number: 3; type: "Journal Article" FEATURES Location/Qualifiers source 1..10878 /mol_type="other DNA" /organism="synthetic DNA construct" promoter 2..937 /label=AOX1 promoter /note="inducible promoter, regulated by methanol" CDS 949..1215 /label=alpha-factor secretion signal /note="N-terminal secretion signal from S. cerevisiae alpha-factor" CDS 1219..3141 /codon_start=1 /note="unnamed protein product; mature TS" /protein_id="SJL88089.1" /translation="MLAPGSSRVELFKRKNSTVPFEDKAGKVTERVVHSFRLPALVNVD GVMVAIADARYDTSNDNSLIDTVAKYSVDDGETWETQIAIKNSRVSSVSRVVDPTVIVK GNKLYVLVGSYYSSRSYWSSHGDARDWDILLAVGEVTKSIAGGKITASIKWGSPVSLKK FFPAEMEGMHTNQFLGGAGVAIVASNGNLVYPVQVTNKRKQVFSKIFYSEDDGKTWKFG KGRSDFGCSEPVALEWEGKLIINTRVDWKRRLVYESSDMGNTWVEAVGTLSRVWGPSPK SDHPGSQSSFTAVTIEGMRVMLFTHPLNFKGRWLRDRLNLWLTDNQRIYNVGQVSIGDE NSAYSSVLYKDDKLYCLHEINTDEVYSLVFARLVGELRIIKSVLRSWKNWDSHLSSICT PADPAASSSESGCGPAVTTVGLVGFLSGNASQNVWEDAYRCVNASTANAERVRNGLKFA GVGGGALWPVSQQGQNQRYRFANHAFTLVASVTIHEAPRAASPLLGASLDSSGGKKLLG LSYDEKHQWQPIYGSTPVTPTGSWETGKRYHVVLTVANKIGSVYIDGELLEGSGQTVVP DGRTPDISHFYVGGYGRSDMPTISHVTVNNVLLYNRQLNTEEIRTLFLSQDLIGTEAHM DSSSDT" misc_feature 3151..3189 /label=E-tag /note="E-tag" CDS 3199..4090 /codon_start=1 /note="unnamed protein product; fd g3 fragment" /protein_id="SJL88090.1" /translation="TVESCLAKPHTENSFTNVWKDDKTLDRYANYEGCLWNATGVVVCT GDETQCYGTWVPIGLAIPENEGGGSEGGGSEGGGSEGGGTKPPEYGDTPIPGYTYINPL DGTYPPGTEQNPANPNPSLEESQPLNTFMFQNNRFRNRQGALTVYTGTVTQGTDPVKTY YQYTPVSSKAMYDAYWNGKFRDCAFHSGFNEDPFVCEYQGQSSDLPQPPVNAGGGSGGG SGGGSEGGGSEGGGSEGGGSEGGGSGGGSGSGDFDYEKMANANKGAMTENADENALQSD AKGKLDSVATDYGAAI" terminator 4176..4422 /label=AOX1 terminator /note="transcription terminator for AOX1" CDS complement(4835..7369) /codon_start=1 /gene="Pichia pastoris HIS4" /product="multifunctional enzyme, required for histidine biosynthesis" /label=PpHIS4 /note="auxotrophic marker for Pichia pastoris" /translation="MTFPLLPAYASVAEFDNSLSLVGKAVFPYAADQLHNLIKFTQSTE LQVNVQVESSVTEDQFEELIDNLLKLYNNGINEVILDLDLAERVVQRMIPGARVIYRTL VDKVASLPANASIAVPFSSPLGDLKSFTNGGSRTVYAFSETAKLVDVTSTVASGIIPII DARQLTTEYELSEDVKKFPVSEILLASLTTDRPDGLFTTLVADSSNYSLGLVYSSKKSI PEAIRTQTGVYQSRRHGLWYKGATSGATQKLLGIELDCDGDCLKFVVEQTGVGFCHLER TSCFGQSKGLRAMEATLWDRKSNAPEGSYTKRLFDDEVLLNAKIREEADELAEAKSKED IAWECADLFYFALVRCAKYGVTLDEVERNLDMKSLKVTRRKGDAKPGYTKEQPKEESKP KEVPSEGRIELCKIDVSKASSQEIEDALRRPIQKTEQIMELVKPIVDNVRQNGDKALLE LTAKFDGVALKTPVLEAPFPEELMQLPDNVKRAIDLSIDNVRKFHEAQLTETLQVETCP GVVCSRFARPIEKVGLYIPGGTAILPSTSLMLGVPAKVAGCKEIVFASPPKKDGTLTPE VIYVAHKVGAKCIVLAGGAQAVAAMAYGTETVPKCDKIFGPGNQFVTAAKMMVQNDTSA LCSIDMPAGPSEVLVIADKYADPDFVASDLLSQAEHGIDSQVILLAVDMTDKELARIED AVHNQAVQLPRVEIVRKCIAHSTTLSVATYEQALEMSNQYAPEHLILQIENASSYVDQV QHAGSVFVGAYSPESCGDYSSGTNHTLPTYGYARQYSGVNTATFQKFITSQDVTPEGLK HIGQAVMDLAAVEGLDAHRNAVKVRMEKLGLI" CDS complement(4838..7369) /codon_start=1 /note="unnamed protein product; Pichia HIS4" /protein_id="SJL88091.1" /translation="MTFPLLPAYASVAEFDNSLSLVGKAVFPYAADQLHNLIKFTQSTE LQVNVQVESSVTEDQFEELIDNLLKLYNNGINEVILDLDLAERVVQRMIPGARVIYRTL VDKVASLPANASIAVPFSSPLGDLKSFTNGGSRTVYAFSETAKLVDVTSTVASGIIPII DARQLTTEYELSEDVKKFPVSEILLASLTTDRPDGLFTTLVADSSNYSLGLVYSSKKSI PEAIRTQTGVYQSRRHGLWYKGATSGATQKLLGIELDCDGDCLKFVVEQTGVGFCHLER TSCFGQSKGLRAMEATLWDRKSNAPEGSYTKRLFDDEVLLNAKIREEADELAEAKSKED IAWECADLFYFALVRCAKYGVTLDEVERNLDMKSLKVTRRKGDAKPGYTKEQPKEESKP KEVPSEGRIELCKIDVSKASSQEIEDALRRPIQKTEQIMELVKPIVDNVRQNGDKALLE LTAKFDGVALKTPVLEAPFPEELMQLPDNVKRAIDLSIDNVRKFHEAQLTETLQVETCP GVVCSRFARPIEKVGLYIPGGTAILPSTSLMLGVPAKVAGCKEIVFASPPKKDGTLTPE VIYVAHKVGAKCIVLAGGAQAVAAMAYGTETVPKCDKIFGPGNQFVTAAKMMVQNDTSA LCSIDMPAGPSEVLVIADKYADPDFVASDLLSQAEHGIDSQVILLAVDMTDKELARIED AVHNQAVQLPRVEIVRKCIAHSTTLSVATYEQALEMSNQYAPEHLILQIENASSYVDQV QHAGSVFVGAYSPESCGDYSSGTNHTLPTYGYARQYSGVNTATFQKFITSQDVTPEGLK HIGQAVMDLAAVEGLDAHRNAVKVRMEKLGLI" misc_feature 7724..8480 /label=AOX1 3' fragment /note="region downstream of Pichia pastoris AOX1 gene" misc_feature 8623..8763 /label=bom /note="basis of mobility region from pBR322" rep_origin complement(8949..9537) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(9711..10568) /label=AmpR /note="beta-lactamase" promoter complement(10569..10673) /label=AmpR promoter