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:
- pAV012
- 载体抗性:
- Ampicillin
- 载体长度:
- 4577 bp
- 载体类型:
- Cloning vector
- 复制子:
- R6K γ ori
- 载体来源:
- White KP, Victorsen A.
- 启动子:
- mPGK
pAV012 载体载体图谱
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。
pAV012 载体载体序列
LOCUS 40924_5089 4577 bp DNA circular SYN 17-DEC-2018 DEFINITION Cloning vector pAV012, complete sequence. ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4577) AUTHORS White KP, Victorsen A. TITLE Direct Submission JOURNAL Submitted (15-JUL-2013) Human Genetics, The University of Chicago, 5801 South Ellis Avenue, Chicago, IL 60637, USA REFERENCE 2 (bases 1 to 4577) TITLE Direct Submission REFERENCE 3 (bases 1 to 4577) AUTHORS . TITLE Direct Submission COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Submitted (15-JUL-2013) Human Genetics, The University of Chicago, 5801 South Ellis Avenue, Chicago, IL 60637, USA" COMMENT SGRef: number: 2; type: "Journal Article" FEATURES Location/Qualifiers source 1..4577 /mol_type="other DNA" /organism="synthetic DNA construct" rep_origin 26..414 /label=R6K gamma ori /note="gamma replication origin from E. coli plasmid R6K; requires the R6K initiator protein pi for replication" promoter 509..613 /label=AmpR promoter CDS 614..1471 /label=AmpR /note="beta-lactamase" misc_feature 1583..1867 /note="sGFP exon 1; 1st exon of superfolder green fluorescent protein" primer_bind 1583..1604 /label=forward recombineering primer binding site /note="forward recombineering primer binding site" misc_feature 1941..1974 /label=loxP /note="loxP" protein_bind 1941..1974 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." promoter 1989..2488 /label=PGK promoter /note="mouse phosphoglycerate kinase 1 promoter" promoter 2505..2552 /label=EM7 promoter /note="synthetic bacterial promoter" CDS 2571..3371 /label=NeoR/KanR /note="aminoglycoside phosphotransferase from Tn5" protein_bind complement(3507..3540) /label=loxP /note="Cre-mediated recombination occurs in the 8-bp core sequence (ATGTATGC) (Shaw et al., 2021)." promoter 3604..3622 /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" CDS 4109..4126 /label=tetracysteine tag /note="tetracysteine peptide that binds biarsenical labeling reagents" CDS 4148..4171 /label=Strep-Tag II /note="peptide that binds Strep-Tactin(R), an engineered form of streptavidin" CDS 4172..4192 /label=TEV site /note="tobacco etch virus (TEV) protease recognition and cleavage site" CDS 4235..4258 /label=FLAG /note="FLAG(R) epitope tag, followed by an enterokinase cleavage site" primer_bind complement(4265..4288) /label=reverse recombineering primer binding site /note="reverse recombineering primer binding site" oriT complement(4334..4443) /direction=LEFT /label=oriT /note="incP origin of transfer"