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 pEPI-1 vector contains two mammalian transcription units and a total of 305 CpG islands, which are located predominantly within the vector elements necessary for bacterial propagation and known to be counterproductive for persistent long-term transgene expression. The episomal replication of vector pEPI-1 depends on a transcription unit starting from the constitutively expressed Cytomegalovirus immediate early promoter (CMV-IEP) and directed into a 2000 bp long matrix attachment region sequence (MARS) derived from the human β-interferon gene.
- 载体名称:
- pEPI-1
- 载体抗性:
- Kanamycin
- 载体长度:
- 6944 bp
- 载体类型:
- Cloning vector
- 复制子:
- ori
- 载体来源:
- Zhang X, Wang XY, Jia YL, Guo X, Wang YF, Wang TY.
- 感受态:
- DH10b
- 培养温度:
- 37℃
pEPI-1 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。
pEPI-1 载体序列
LOCUS Exported 6944 bp DNA circular SYN 29-AUG-2024
DEFINITION synthetic circular DNA.
ACCESSION .
VERSION .
KEYWORDS pEPI-1
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6944)
TITLE Direct Submission
REFERENCE 2 (bases 1 to 6944)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..6944
/mol_type="other DNA"
/organism="synthetic DNA construct"
enhancer 1..304
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 305..508
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
CDS 553..1269
/label=EGFP
/note="enhanced GFP"
misc_feature 1276..3476
/label=S/MAR
/note="S/MAR"
/note="S/MARs-element(Scaffold or Matrix Attachment
Regions) are typically 70% A/T-rich sequences and are often
found in association with chromosomal origins of
bidirectional replication. It could be used to construct
non-viral episomal vectors in mammalian cells."
misc_feature 3483..3548
/label=MCS
/note="multiple cloning site"
polyA_signal 3672..3793
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
rep_origin complement(3800..4255)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 4282..4386
/label=AmpR promoter
promoter 4388..4745
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
CDS 4780..5571
/label=NeoR/KanR
/note="aminoglycoside phosphotransferase"
polyA_signal 5806..5853
/label=HSV TK poly(A) signal
/note="herpes simplex virus thymidine kinase
polyadenylation signal (Cole and Stacy, 1985)"
rep_origin 6182..6770
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
promoter 6885..6944
/label=CMV
/note="Human cytomegalovirus immediate early
enhancer/promoter"