质粒编号
质粒名称
规格
价格
V010351
pTRIPZ
5 μg
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。
The Thermo Scientific TRIPZ Inducible Lentiviral shRNA Collection was developed in collaboration with Dr. Greg Hannon of Cold Spring Harbor Laboratory (CSHL) and Dr. Steve Elledge of Harvard Medical School. This collection combines the design advantages of microRNA-adapted short hairpin RNA (shRNA) with the pTRIPZ lentiviral inducible vector to create a powerful RNAi trigger capable of producing gene silencing. The vector is engineered to be Tet-On and produces tightly regulated induction of shRNA expression in the presence of doxycycline.
- 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
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"