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:
- pEarleyGate 205
- 载体抗性:
- Chloramphenicol
- 载体长度:
- 12188 bp
- 载体类型:
- Plant Vectors
- 复制子:
- ori
- 载体来源:
- Earley KW, Haag JR, Pontes O, Opper K, Juehne T, Song K, Pikaard CS.
- 拷贝数:
- High copy number
- 启动子:
- MAS
pEarleyGate 205 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。
pEarleyGate 205 载体序列
LOCUS pEarleyGate_205. 12188 bp DNA circular SYN 01-JAN-1980 DEFINITION Gateway(R)-compatible plant transformation vector with a C-terminal TAP tag. ACCESSION . VERSION . KEYWORDS pEarleyGate 205 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 12188) AUTHORS Earley KW, Haag JR, Pontes O, Opper K, Juehne T, Song K, Pikaard CS. TITLE Gateway-compatible vectors for plant functional genomics and proteomics. JOURNAL Plant J. 2006;45:616-29. PUBMED 16441352 REFERENCE 2 (bases 1 to 12188) AUTHORS Pikaard Lab TITLE Direct Submission REFERENCE 3 (bases 1 to 12188) AUTHORS . TITLE Direct Submission COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Plant J."; date: "2006"; volume: "45"; pages: "616-29" COMMENT SGRef: number: 2; type: "Journal Article" COMMENT The sequence was corrected by inserting a G at position 8296. FEATURES Location/Qualifiers source 1..12188 /mol_type="other DNA" /organism="synthetic DNA construct" misc_feature 1..25 /label=LB T-DNA repeat /note="left border repeat from nopaline C58 T-DNA" terminator complement(111..363) /label=MAS terminator /note="mannopine synthase terminator" CDS complement(376..924) /label=BlpR /note="phosphinothricin acetyltransferase" promoter complement(930..1310) /label=MAS promoter /note="mannopine synthase promoter (Velten et al., 1984)" promoter 2306..2651 /label=CaMV 35S promoter /note="strong constitutive promoter from cauliflower mosaic virus" protein_bind 2663..2782 /label=attR1 /note="recombination site for the Gateway(R) LR reaction" promoter 2812..2842 /label=lac UV5 promoter /note="E. coli lac promoter with an 'up' mutation" CDS 2896..3552 /label=CmR /note="chloramphenicol acetyltransferase" CDS 3897..4199 /label=ccdB /note="CcdB, a bacterial toxin that poisons DNA gyrase" protein_bind complement(4243..4367) /label=attR2 /note="recombination site for the Gateway(R) LR reaction" CDS 4381..4458 /label=CBP /note="calmodulin-binding peptide" CDS 4486..4506 /label=TEV site /note="tobacco etch virus (TEV) protease recognition and cleavage site" CDS 4546..4719 /label=ProtA /note="IgG-binding unit of Staphylococcus aureus protein A" CDS 4720..4893 /codon_start=1 /product="IgG-binding unit of Staphylococcus aureus protein A" /label=IgG-binding unit of Staphylococcus aureus prote /note="ProtA" /translation="VDNKFNKEQQNAFYEILHLPNLNEEQRNAFIQSLKDDPSQSANLL AEAKKLNGAQAPK" terminator 4959..5666 /label=OCS terminator /note="octopine synthase terminator" misc_feature 5910..5934 /label=RB T-DNA repeat /note="right border repeat from nopaline C58 T-DNA" CDS 7234..7860 /label=pVS1 StaA /note="stability protein from the Pseudomonas plasmid pVS1 (Heeb et al., 2000)" CDS 8292..9362 /label=pVS1 RepA /note="replication protein from the Pseudomonas plasmid pVS1 (Heeb et al., 2000)" rep_origin 9431..9625 /label=pVS1 oriV /note="origin of replication for the Pseudomonas plasmid pVS1 (Heeb et al., 2000)" misc_feature 9969..10109 /label=bom /note="bom" /note="basis of mobility region from pBR322" rep_origin complement(10295..10883) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(10973..11764) /label=KanR /note="aminoglycoside phosphotransferase"