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:
pLJM1-EGFP
载体抗性:
Ampicillin
载体长度:
8083 bp
载体类型:
Mammalian Expression, Lentiviral
复制子:
ori
筛选标记:
Puromycin
拷贝数:
High Copy
启动子:
hPGK
克隆方法:
Restriction Enzyme
5'引物:
CMV-F
感受态:
DH10B
培养温度:
37℃

pLJM1-EGFP 载体图谱

pLJM1-EGFP8083 bp400800120016002000240028003200360040004400480052005600600064006800720076008000CMV promoterEGFPcPPT/CTShPGK promoterPuroR3' LTR (Delta-U3)SV40 poly(A) signalSV40 oriT7 promoterM13 fwdf1 oriAmpR promoterAmpRoriL4440CAP binding sitelac promoterlac operatorM13 revT3 promoterRSV promoter5' LTR (truncated)HIV-1 PsiRREgp41 peptideProtein Tat

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。

pLJM1-EGFP 载体序列

LOCUS       V001188                 8083 bp    DNA     circular SYN 13-MAY-2021
DEFINITION  Exported.
ACCESSION   V001188
VERSION     V001188
KEYWORDS    pLJM1-EGFP
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 8083)
  AUTHORS   Sancak Y, Peterson TR, Shaul YD, Lindquist RA, Thoreen CC, Bar-Peled
            L, Sabatini DM
  TITLE     The Rag GTPases bind raptor and mediate amino acid signaling to
            mTORC1.
  JOURNAL   Science. 2008 Jun 13. 320(5882):1496-501.
   PUBMED   18497260
REFERENCE   2  (bases 1 to 8083)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 8083)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Science.
            2008 Jun 13. 320(5882):1496-501."
            SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..8083
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        290..493
                     /label="CMV promoter"
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     CDS             538..1254
                     /label="EGFP"
                     /note="enhanced GFP"
     misc_feature    1325..1442
                     /label="cPPT/CTS"
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
     promoter        1491..2001
                     /label="hPGK promoter"
                     /note="human phosphoglycerate kinase 1 promoter"
     CDS             2023..2619
                     /label="PuroR"
                     /note="puromycin N-acetyltransferase"
     LTR             2750..2983
                     /label="3' LTR (Delta-U3)"
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    3055..3189
                     /label="SV40 poly(A) signal"
                     /note="SV40 polyadenylation signal"
     rep_origin      3216..3351
                     /label="SV40 ori"
                     /note="SV40 origin of replication"
     promoter        complement(3372..3390)
                     /label="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3400..3416)
                     /label="M13 fwd"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     rep_origin      3558..4013
                     /label="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        4039..4143
                     /label="AmpR promoter"
     CDS             4144..5001
                     /label="AmpR"
                     /note="beta-lactamase"
     rep_origin      5175..5763
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     primer_bind     5917..5934
                     /label="L4440"
                     /note="L4440 vector, forward primer"
     protein_bind    6051..6072
                     /label="CAP binding site"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        6087..6117
                     /label="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    6125..6141
                     /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)."
     primer_bind     6149..6165
                     /label="M13 rev"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     promoter        6186..6204
                     /label="T3 promoter"
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        6232..6458
                     /label="RSV promoter"
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             6459..6639
                     /label="5' LTR (truncated)"
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    6686..6811
                     /label="HIV-1 Psi"
                     /note="packaging signal of human immunodeficiency virus
                     type 1"
     misc_feature    7304..7537
                     /label="RRE"
                     /note="The Rev response element (RRE) of HIV-1 allows for
                     Rev-dependent mRNA export from the nucleus to the
                     cytoplasm."
     CDS             7722..7766
                     /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             7915..7956
                     /note="Protein Tat from Human immunodeficiency virus type 1
                     group M subtype B (isolate WMJ22). Accession#: P12509"
                     /label="Protein Tat"