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。
pCMV5 is a Eukaryotic expression vector.
- 载体名称:
- pCMV5
- 载体抗性:
- Ampicillin
- 载体长度:
- 4657 bp
- 载体类型:
- Eukaryotic expression vector
- 复制子:
- ori
- 载体来源:
- Andersson S, Davis DL, Dahlback H, Jornvall H, Russell DW.
- 启动子:
- CMV
- 感受态:
- DH10B
- 培养温度:
- 37℃
pCMV5 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。
pCMV5 载体序列
LOCUS 40924_12030 4657 bp DNA circular SYN 17-DEC-2018 DEFINITION Eukaryotic expression vector pCMV5, complete sequence. ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4657) AUTHORS Andersson S, Davis DL, Dahlback H, Jornvall H, Russell DW. TITLE Cloning, structure, and expression of the mitochondrial cytochrome P-450 sterol 26-hydroxylase, a bile acid biosynthetic enzyme JOURNAL J. Biol. Chem. 264 (14), 8222-8229 (1989) PUBMED 2722778 REFERENCE 2 (bases 1 to 4657) AUTHORS Andersson S, Davis DL, Dahlback H, Jornvall H, Russell DW. TITLE Direct Submission JOURNAL Submitted (25-FEB-2000) Molecular Genetics, University of Texas Southwestern Medical Center at Dallas, 5323 Harry Hines Blvd., Dallas, TX 75235, USA REFERENCE 3 (bases 1 to 4657) TITLE Direct Submission REFERENCE 4 (bases 1 to 4657) AUTHORS . TITLE Direct Submission COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "J. Biol. Chem."; date: "1989"; volume: "264"; issue: "14"; pages: "8222-8229" COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted (25-FEB-2000) Molecular Genetics, University of Texas Southwestern Medical Center at Dallas, 5323 Harry Hines Blvd., Dallas, TX 75235, USA" COMMENT SGRef: number: 3; type: "Journal Article" FEATURES Location/Qualifiers source 1..4657 /mol_type="other DNA" /organism="synthetic DNA construct" primer_bind 142..158 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" enhancer 319..698 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 699..902 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" polyA_signal 987..1609 /label=hGH poly(A) signal /note="human growth hormone polyadenylation signal" promoter 1636..1966 /label=SV40 promoter /note="SV40 enhancer and early promoter" promoter complement(2005..2023) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(2038..2054) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" protein_bind complement(2062..2078) /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 complement(2086..2116) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind complement(2131..2152) /label=CAP binding site /note="CAP binding activates transcription in the presence of cAMP." rep_origin complement(2439..3027) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(3201..4058) /codon_start=1 /label=AmpR /note="beta-lactamase" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(4059..4163) /label=AmpR promoter rep_origin 4190..4618 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis"