pLV3-CMV-MCS-6×His-Neo 载体 (V018098)

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:
pLV3-CMV-MCS-6×His-Neo
载体抗性:
Ampicillin
载体长度:
7606 bp
载体类型:
Protein expression
复制子:
ori
宿主:
Mammalian cells, Lentivirus
筛选标记:
Neo/G418
启动子:
CMV
5'引物:
CMV-F
感受态:
Stbl3
培养温度:
37℃

pLV3-CMV-MCS-6×His-Neo 载体图谱

pLV3-CMV-MCS-6×His-Neo7606 bp3006009001200150018002100240027003000330036003900420045004800510054005700600063006600690072007500RSV promoterHIV-1 ΨRREgp41 peptideProtein TatcPPT/CTSCMV promoter6xHisEF-1α core promoter5' LTR (truncated)NeoR/KanRWPRE3' LTR (ΔU3)SV40 poly(A) signalSV40 oriM13 revlac operatorlac promoterCAP binding siteoriAmpRAmpR promoterM13 fwd

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。

pLV3-CMV-MCS-6×His-Neo 载体序列

LOCUS       .                       7606 bp    DNA     circular UNK 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   
VERSION     
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     promoter        6..233
                     /label="RSV promoter"
                     /note="Rous sarcoma virus enhancer/promoter"
     misc_feature    458..583
                     /label="HIV-1 Ψ"
                     /note="packaging signal of human immunodeficiency virus
                     type 1"
     misc_feature    1076..1309
                     /label="RRE"
                     /note="The Rev response element (RRE) of HIV-1 allows for
                     Rev-dependent mRNA export from the nucleus to the
                     cytoplasm."
     CDS             1493..1537
                     /label="gp41 peptide"
                     /note="antigenic peptide corresponding to amino acids 655
                     to 669 of the HIV envelope protein gp41 (Lutje Hulsik et
                     al., 2013)"
     CDS             1686..1727
                     /gene="Protein Tat"
                     /label="Protein Tat"
                     /note="Protein Tat from Human immunodeficiency virus type 1
                     group M subtype B (isolate WMJ22). Accession#: P12509"
     misc_feature    1804..1920
                     /label="cPPT/CTS"
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
     promoter        2010..2213
                     /label="CMV promoter"
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     CDS             2308..2325
                     /label="6xHis"
                     /note="6xHis affinity tag"
     promoter        2375..2586
                     /label="EF-1α core promoter"
                     /note="core promoter for human elongation factor EF-1α"
     LTR             2599..2867
                     /label="5' LTR (truncated)"
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
     CDS             2900..3691
                     /label="NeoR/KanR"
                     /note="aminoglycoside phosphotransferase"
     misc_feature    3701..4289
                     /label="WPRE"
                     /note="woodchuck hepatitis virus posttranscriptional
                     regulatory element"
     LTR             4363..4596
                     /label="3' LTR (ΔU3)"
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4668..4802
                     /label="SV40 poly(A) signal"
                     /note="SV40 polyadenylation signal"
     rep_origin      4808..4943
                     /label="SV40 ori"
                     /note="SV40 origin of replication"
     primer_bind     complement(4981..4997)
                     /label="M13 rev"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     protein_bind    complement(5005..5021)
                     /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-β-D-thiogalactopyranoside (IPTG)."
     promoter        complement(5029..5059)
                     /label="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(5074..5095)
                     /label="CAP binding site"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(5383..5971)
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     CDS             complement(6145..7002)
                     /label="AmpR"
                     /note="β-lactamase"
     promoter        complement(7003..7107)
                     /label="AmpR promoter"
     primer_bind     7581..7597
                     /label="M13 fwd"
                     /note="common sequencing primer, one of multiple similar
                     variants"