pIEx/Bac-1 载体图谱和序列

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。

The dual-purpose pIEx/Bac vectors are designed for cloning and high-level expression of proteins by transiently transfecting Spodoptera-derived insect cells or by generating baculovirus recombinants. Transient transfection and early baculovirus expression is driven by a promoter/enhancer combination that recruits endogenous insect cell transcription machinery, the AcNPV derived hr5 enhancer and ie1 promoter. Late/very late expression in the baculovirus mode is driven by the strong p10 promoter. The pIEx/Bac-1 vector carries an N-terminal Strep•Tag II coding sequence (1) followed by a recognition site for enterokinase.The multiple cloning region is followed by an optional C-terminal His•Tag coding sequence. The presence of two 'gentle elution' tags at both the N- and C-terminus is ideal for dual purification strategies designed to isolate full-length fusion proteins (2). Unique restriction sites are shown on the circle map.

载体名称:: pIEx/Bac-1

载体抗性:: Ampicillin

载体长度:: 6796 bp

载体类型:: Insect Cell Vectors

复制子:: ori

载体来源:: Novagen (EMD Millipore)

拷贝数:: High copy number

启动子:: IE1

克隆方法:: Enzyme digestion and ligation

载体标签:: C-His,N-Strep

pIEx/Bac-1 载体图谱

复制序列 NovoBuilder中查看图谱

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。

pIEx/Bac-1 载体序列

下载GenBank文件(.gb)

LOCUS       pIEx_Bac-1.        6796 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Insect cell and baculovirus vector encoding a cleavable N-terminal 
            Strep-Tag(R) II and a C-terminal 10xHis tag.
ACCESSION   .
VERSION     .
KEYWORDS    pIEx Bac-1
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6796)
  AUTHORS   Novagen (EMD Millipore)
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 6796)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6796
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_recomb     329..1156
                     /label=baculovirus recombination region (lef2/ORF603)
                     /note="contains ORF603 and part of lef2"
     enhancer        1196..1678
                     /label=hr5 enhancer
                     /note="baculovirus early transcription enhancer"
     promoter        1682..2273
                     /label=IE1 promoter
                     /note="promoter of the ie1 gene from the baculovirus
                     Autographa californica"
     promoter        2283..2392
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     CDS             2394..2396
                     /codon_start=1
                     /product="start codon"
                     /label=start codon
                     /note="ATG"
                     /translation="M"
     CDS             2403..2426
                     /label=Strep-Tag II
                     /note="peptide that binds Strep-Tactin(R), an engineered 
                     form
                     of streptavidin"
     CDS             2433..2447
                     /label=enterokinase site
                     /note="enterokinase recognition and cleavage site"
     misc_feature    2447..2506
                     /label=MCS
                     /note="MCS"
                     /note="multiple cloning site"
     CDS             2517..2543
                     /label=9xHis
                     /note="9xHis affinity tag"
     terminator      2565..2871
                     /label=IE1 terminator
                     /note="terminator of the ie1 gene from the baculovirus 
                     Autographa californica"
     misc_recomb     2900..4257
                     /label=baculovirus recombination region (ORF1629)
                     /note="contains part of ORF1629"
     rep_origin      complement(4520..5108)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(5282..6139)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(6140..6244)
                     /label=AmpR promoter
     rep_origin      6271..6726
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"

pIEx/Bac-1 载体 (V011638)

pIEx/Bac-1 载体图谱

质粒操作方法

pIEx/Bac-1 载体序列