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:
- pOPINE
- 载体抗性:
- Ampicillin
- 载体长度:
- 5534 bp
- 载体类型:
- Expression vector
- 复制子:
- ori
- 载体来源:
- Berrow NS, Alderton D, Sainsbury S, Nettleship J, Assenberg R, Rahman N, Stuart DI, Owens RJ.
- 启动子:
- chicken β-actin
pOPINE 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。
pOPINE 载体序列
LOCUS 40924_33767 5534 bp DNA circular SYN 18-DEC-2018
DEFINITION Expression vector pOPINE, complete sequence.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5534)
AUTHORS Berrow NS, Alderton D, Sainsbury S, Nettleship J, Assenberg R,
Rahman N, Stuart DI, Owens RJ.
TITLE A versatile ligation-independent cloning method suitable for
high-throughput expression screening applications
JOURNAL Nucleic Acids Res. 35 (6), E45 (2007)
PUBMED 17317681
REFERENCE 2 (bases 1 to 5534)
AUTHORS Berrow NS.
TITLE Direct Submission
JOURNAL Submitted (17-JAN-2007) Oxford Protein Production Facility, Oxford
University, Roosevelt Drive, Oxford, Oxfordshire OX3 7BN, U.K.
REFERENCE 3 (bases 1 to 5534)
TITLE Direct Submission
REFERENCE 4 (bases 1 to 5534)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Nucleic
Acids Res."; date: "2007"; volume: "35"; issue: "6"; pages: "E45"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
(17-JAN-2007) Oxford Protein Production Facility, Oxford University,
Roosevelt Drive, Oxford, Oxfordshire OX3 7BN, U.K."
COMMENT SGRef: number: 3; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..5534
/mol_type="other DNA"
/organism="synthetic DNA construct"
misc_recomb 181..1008
/label=baculovirus recombination region (lef2/ORF603)
/note="contains ORF603 and part of lef2"
enhancer 1140..1443
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 1449..1726
/label=chicken beta-actin promoter
promoter 2150..2168
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 2169..2193
/label=lac operator
/note="The lac repressor binds to the lac operator to
inhibit transcription in E. coli. This inhibition can be
relieved by adding lactose or
isopropyl-beta-D-thiogalactopyranoside (IPTG)."
promoter 2209..2318
/label=p10 promoter
/note="baculovirus promoter for expression in insect cells"
regulatory 2319..2326
/regulatory_class="ribosome_binding_site"
misc_feature 2321..2335
/label=5' in-fusion site (after NcoI digest)
/note="5' in-fusion site (after NcoI digest)"
CDS 2339..2356
/label=6xHis
/note="6xHis affinity tag"
protein_bind 2409..2430
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 2445..2475
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 2483..2499
/label=lac operator
/bound_moiety="lac repressor encoded by lacI"
/note="The lac repressor binds to the lac operator to
inhibit transcription in E. coli. This inhibition can be
relieved by adding lactose or
isopropyl-beta-D-thiogalactopyranoside (IPTG)."
CDS 2519..2692
/label=lacZ-alpha
/note="LacZ-alpha fragment of beta-galactosidase"
CDS 2722..2739
/label=6xHis
/note="6xHis affinity tag"
polyA_signal 2894..2949
/label=beta-globin poly(A) signal
/note="rabbit beta-globin polyadenylation signal (Gil and
Proudfoot, 1987)"
terminator 3037..3084
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
misc_recomb 3097..3802
/label=baculovirus recombination region (ORF1629)
/note="contains part of ORF1629"
protein_bind complement(3811..3844)
/label=loxP
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (ATGTATGC) (Shaw et al., 2021)."
rep_origin complement(4012..4599)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(4615..5472)
/label=AmpR
/note="beta-lactamase"