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:
- pNL4-3
- 载体抗性:
- Ampicillin
- 载体长度:
- 14825 bp
- 载体类型:
- HIV-1 vector
- 复制子:
- ori
- 载体来源:
- Adachi A, Gendelman HE, Koenig S, Folks T, Willey R, Rabson A, Martin MA.
- 感受态:
- DH5alpha
- 培养温度:
- 37℃
pNL4-3 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。
pNL4-3 载体序列
LOCUS V004299 14825 bp DNA circular SYN 18-DEC-2018 DEFINITION Exported. ACCESSION V004299 VERSION V004299 KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct . REFERENCE 1 (bases 1 to 14825) AUTHORS Adachi A, Gendelman HE, Koenig S, Folks T, Willey R, Rabson A, Martin MA. TITLE Production of acquired immunodeficiency syndrome-associated retrovirus in human and nonhuman cells transfected with an infectious molecular clone JOURNAL J. Virol. 59 (2), 284-291 (1986) PUBMED 3016298 REFERENCE 2 (bases 1 to 14825) AUTHORS Bosche WJ, Poon DTK., Ott DE, Hu W-S., Gorelick RJ. TITLE Complete Plasmid Sequence of pNL4-3 JOURNAL Unpublished REFERENCE 3 (bases 1 to 14825) AUTHORS Bosche WJ, Poon DTK., Ott DE, Hu W-S., Gorelick RJ. TITLE Direct Submission JOURNAL Submitted (28-NOV-2000) AIDS Vaccine Program, SAIC Frederick and Drug Resistance Program, National Cancer Institute at Frederick, PO Box B, Frederick, Maryland 21702-1202, USA REFERENCE 4 (bases 1 to 14825) AUTHORS Strebel KJ, Martin MA. TITLE Direct Submission JOURNAL Submitted (24-MAY-2010) NIAID, Viral Biochemistry, National Institutes of Health, Bethesda, MD, USA REFERENCE 5 (bases 1 to 14825) TITLE Direct Submission REFERENCE 6 (bases 1 to 14825) AUTHORS . TITLE Direct Submission COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "J. Virol."; date: "1986"; volume: "59"; issue: "2"; pages: "284-291" SGRef: number: 2; type: "Journal Article"; journalName: "Unpublished" SGRef: number: 3; type: "Journal Article"; journalName: "Submitted (28-NOV-2000) AIDS Vaccine Program, SAIC Frederick and Drug Resistance Program, National Cancer Institute at Frederick, PO Box B, Frederick, Maryland 21702-1202, USA" SGRef: number: 4; type: "Journal Article"; journalName: "Submitted (24-MAY-2010) NIAID, Viral Biochemistry, National Institutes of Health, Bethesda, MD, USA" SGRef: number: 5; type: "Journal Article" On May 24, 2010 this sequence version replaced AF324493.1. FEATURES Location/Qualifiers source 1..14825 /mol_type="other DNA" /organism="synthetic DNA construct" repeat_region 1..634 /label="5' long terminal repeat" /note="5' long terminal repeat" LTR 377..634 /label="5' LTR (truncated)" /note="truncated 5' long terminal repeat (LTR) from HIV-1" repeat_region 454..550 /note="R; 5' copy" misc_feature 681..806 /label="HIV-1 Psi" /note="packaging signal of human immunodeficiency virus type 1" CDS 790..2289 /label="HIV-1 gag" /note="gag protein from human immunodeficiency virus 1" CDS 2085..5093 /label="HIV-1 pol" /note="pol protein from human immunodeficiency virus 1" CDS 5041..5616 /gene="vif" /label="Virion infectivity factor" /note="Virion infectivity factor from Human immunodeficiency virus type 1 group M subtype B (isolate NY5). Accession#: P12504" CDS 5559..5849 /codon_start=1 /product="vpr protein" /label="vpr protein" /protein_id="AAK08485.1" /translation="MEQAPEDQGPQREPYNEWTLELLEELKSEAVRHFPRIWLHNLGQH IYETYGDTWAGVEAIIRILQQLLFIHFRIGCRHSRIGVTRQRRARNGASRS" misc_feature 5743..5748 /label="EcoRI site of recombination" /note="EcoRI site of recombination" CDS 5830..6003 /note="Protein Tat from Human immunodeficiency virus type 1 group M subtype B (isolate BH5). Accession#: P04612" /label="Protein Tat" CDS 6221..8779 /gene="env" /label="Envelope glycoprotein gp160" /note="Envelope glycoprotein gp160 from Human immunodeficiency virus type 1 group M subtype B (isolate MFA). Accession#: P19551" CDS 8787..8813 /gene="nef" /label="Protein Nef" /note="Protein Nef from Human immunodeficiency virus type 1 group M subtype D (isolate Z84). Accession#: P12481" LTR 9076..9709 /label="3' LTR" /note="3' long terminal repeat (LTR) from HIV-1" misc_feature 9710..11283 /label="3' cellular genomic sequence" /note="3' cellular genomic sequence" rep_origin complement(11520..12108) /direction=LEFT /label="ori" /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(12282..13139) /label="AmpR" /note="beta-lactamase" promoter complement(13140..13244) /label="AmpR promoter" misc_feature 13648..14825 /label="5' cellular genomic sequence" /note="5' cellular genomic sequence"