pTRIPZ 载体 (V010351)

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:
pTRIPZ
载体抗性:
Ampicillin
载体长度:
13320 bp
载体类型:
Viral Expression & Packaging Vectors
复制子:
ori
载体来源:
Open Biosystems
拷贝数:
High copy number
启动子:
UbC

pTRIPZ 载体图谱

pTRIPZ13320 bp6001200180024003000360042004800540060006600720078008400900096001020010800114001200012600132003' LTRHIV-1 PsiRREgp41 peptideProtein TatcPPT/CTSBleoREM7 promotertet operatortet operatortet operatortet operatortet operatortet operatorminimal CMV promoterTurboRFP5' miR-30a3' miR-30aT7 promoterUbC promoterloxP511rtTA3loxP511PuroRWPRE3' LTR (Delta-U3)bGH poly(A) signalf1 oriSV40 promoterHygR (inactivated)SV40 poly(A) signallac promoterCAP binding siteoriAmpRAmpR promoterSV40 poly(A) signal

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。

pTRIPZ 载体序列

LOCUS       V010351                13320 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  Exported.
ACCESSION   V010351
VERSION     V010351
KEYWORDS    pTRIPZ
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 13320)
  AUTHORS   Open Biosystems
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 13320)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
            Excision of rtTA3 using Cre recombinase or BamHI yields a vector
            that can be used for Tet-repressible expression.
FEATURES             Location/Qualifiers
     source          1..13320
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..634
                     /label="3' LTR"
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    681..806
                     /label="HIV-1 Psi"
                     /note="packaging signal of human immunodeficiency virus
                     type 1"
     misc_feature    1303..1536
                     /label="RRE"
                     /note="The Rev response element (RRE) of HIV-1 allows for
                     Rev-dependent mRNA export from the nucleus to the
                     cytoplasm."
     CDS             1721..1765
                     /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             1914..1955
                     /note="Protein Tat from Human immunodeficiency virus type 1
                     group M subtype B (isolate WMJ22). Accession#: P12509"
                     /label="Protein Tat"
     misc_feature    2063..2180
                     /label="cPPT/CTS"
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
     CDS             complement(2248..2619)
                     /label="BleoR"
                     /note="antibiotic-binding protein"
     promoter        complement(2639..2686)
                     /label="EM7 promoter"
                     /note="synthetic bacterial promoter"
     protein_bind    2722..2740
                     /label="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2757..2775
                     /gene="tetO"
                     /label="tetracycline repressor TetR binding site"
                     /bound_moiety="tetracycline repressor TetR"
                     /note="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2793..2811
                     /gene="tetO"
                     /label="tetracycline repressor TetR binding site"
                     /bound_moiety="tetracycline repressor TetR"
                     /note="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2829..2847
                     /gene="tetO"
                     /label="tetracycline repressor TetR binding site"
                     /bound_moiety="tetracycline repressor TetR"
                     /note="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2864..2882
                     /gene="tetO"
                     /label="tetracycline repressor TetR binding site"
                     /bound_moiety="tetracycline repressor TetR"
                     /note="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2900..2918
                     /gene="tetO"
                     /label="tetracycline repressor TetR binding site"
                     /bound_moiety="tetracycline repressor TetR"
                     /note="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        2930..2968
                     /label="minimal CMV promoter"
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     CDS             3005..3697
                     /label="TurboRFP"
                     /note="red fluorescent protein from Entacmaea quadricolor"
     ncRNA           3713..3806
                     /label="5' miR-30a"
                     /note="sequence upstream of the 71-nt precursor of the
                     human miR-30a microRNA (Zeng et al., 2002)"
     ncRNA           3832..3944
                     /label="3' miR-30a"
                     /note="sequence downstream of the 71-nt precursor of the
                     human miR-30a microRNA (Zeng et al., 2002)"
     promoter        complement(4039..4057)
                     /label="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        4070..5281
                     /label="UbC promoter"
                     /note="human ubiquitin C promoter"
     protein_bind    5294..5327
                     /label="Cre recombinase binding site"
                     /bound_moiety="Cre recombinase"
                     /note="loxP511"
                     /note="Cre-mediated recombination occurs in the 8-bp core
                     sequence (GTATACAT). loxP511 sites are compatible with each
                     other, but incompatible with wild-type loxP sites."
     CDS             5334..6038
                     /label="rtTA3"
                     /note="reverse tetracycline transactivator 3"
     protein_bind    complement(6042..6075)
                     /label="loxP511"
                     /note="Cre-mediated recombination occurs in the 8-bp core
                     sequence (ATGTATAC) (Shaw et al., 2021). loxP511 sites are
                     compatible with each other, but incompatible with wild-type
                     loxP sites."
     misc_feature    6098..6680
                     /label="IRES2"
                     /note="internal ribosome entry site (IRES) of the
                     encephalomyocarditis virus (EMCV)"
     CDS             6679..7272
                     /label="PuroR"
                     /note="puromycin N-acetyltransferase"
     misc_feature    7291..7879
                     /label="WPRE"
                     /note="woodchuck hepatitis virus posttranscriptional
                     regulatory element"
     LTR             8086..8319
                     /label="3' LTR (Delta-U3)"
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    8426..8650
                     /label="bGH poly(A) signal"
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      8696..9124
                     /label="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        9138..9467
                     /label="SV40 promoter"
                     /note="SV40 enhancer and early promoter"
     CDS             9516..10545
                     /label="HygR (inactivated)"
                     /note="HygR (inactivated)"
                     /note="The hygromycin resistance gene was inactivated by
                     destroying an EcoRI site within the coding sequence."
     polyA_signal    10675..10796
                     /label="SV40 poly(A) signal"
                     /note="SV40 polyadenylation signal"
     promoter        complement(10893..10923)
                     /label="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(10938..10959)
                     /label="CAP binding site"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(11247..11835)
                     /direction=LEFT
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     CDS             complement(12009..12866)
                     /label="AmpR"
                     /note="beta-lactamase"
     promoter        complement(12867..12971)
                     /label="AmpR promoter"
     polyA_signal    13019..13153
                     /label="SV40 poly(A) signal"
                     /note="SV40 polyadenylation signal"