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.
| 载体基本信息 | |||
| 载体名称 | pTRIPZ empty | 载体抗性 | Ampicillin |
|---|---|---|---|
| 载体长度 | 13320 bp | 载体类型 | Lentiviral vectors |
| 筛选标记 | Puromycin | 拷贝数 | High copy number |
| 启动子 | TetO | 克隆方法 | Enzyme digestion and ligation |
pTRIPZ empty 载体载体图谱
pTRIPZ empty 载体载体序列
LOCUS Exported 13320 bp ds-DNA circular SYN 13-JAN-2022
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 13320)
TITLE Direct Submission
FEATURES Location/Qualifiers
source 1..13320
/organism="synthetic DNA construct"
/mol_type="other DNA"
LTR 1..634
/note="3 LTR"
/note="3 long terminal repeat (LTR) from HIV-1"
misc_feature 681..806
/note="HIV-1 Psi"
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 1303..1536
/note="RRE"
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
misc_feature 2063..2180
/note="cPPT/CTS"
/note="central polypurine tract and central termination
sequence of HIV-1"
CDS complement(2245..2619)
/codon_start=1
/gene="Sh ble from Streptoalloteichus hindustanus"
/product="antibiotic-binding protein"
/note="BleoR"
/note="confers resistance to bleomycin, phleomycin, and
Zeocin(TM)"
/translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
EFALRDPAGNCVHFVAEEQD"
promoter complement(2629..2647)
/note="T7 promoter"
/note="promoter for bacteriophage T7 RNA polymerase"
promoter complement(2639..2686)
/note="EM7 promoter"
/note="synthetic bacterial promoter "
protein_bind 2722..2740
/gene="tetO"
/bound_moiety="tetracycline repressor TetR"
/note="tet operator"
/note="bacterial operator O2 for the tetR and tetA genes"
protein_bind 2757..2775
/gene="tetO"
/bound_moiety="tetracycline repressor TetR"
/note="tet operator"
/note="bacterial operator O2 for the tetR and tetA genes"
protein_bind 2793..2811
/gene="tetO"
/bound_moiety="tetracycline repressor TetR"
/note="tet operator"
/note="bacterial operator O2 for the tetR and tetA genes"
protein_bind 2829..2847
/gene="tetO"
/bound_moiety="tetracycline repressor TetR"
/note="tet operator"
/note="bacterial operator O2 for the tetR and tetA genes"
protein_bind 2864..2882
/gene="tetO"
/bound_moiety="tetracycline repressor TetR"
/note="tet operator"
/note="bacterial operator O2 for the tetR and tetA genes"
protein_bind 2900..2918
/gene="tetO"
/bound_moiety="tetracycline repressor TetR"
/note="tet operator"
/note="bacterial operator O2 for the tetR and tetA genes"
promoter 2930..2968
/note="minimal CMV promoter"
/note="human cytomegalovirus (CMV) immediate early
promoter"
CDS 3005..3700
/codon_start=1
/product="red fluorescent protein from Entacmaea
quadricolor"
/note="TurboRFP"
/note="mammalian codon-optimized"
/translation="MSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMKIKV
VEGGPLPFAFDILATSFMYGSKAFINHTQGIPDFFKQSFPEGFTWERITTYEDGGVLTA
TQDTSFQNGCIIYNVKINGVNFPSNGPVMQKKTRGWEANTEMLYPADGGLRGHSQMALK
LVGGGYLHCSFKTTYRSKKPAKNLKMPGFHFVDHRLERIKEADKETYVEQHEMAVAKYC
DLPSKLGHR"
ncRNA 3713..3806
/note="5 miR-30a"
/note="sequence upstream of the 71-nt precursor of the
human miR-30a microRNA (Zeng et al., 2002)"
/ncRNA_class="miRNA"
ncRNA 3832..3944
/note="3 miR-30a"
/note="sequence downstream of the 71-nt precursor of the
human miR-30a microRNA (Zeng et al., 2002)"
/ncRNA_class="miRNA"
promoter complement(4039..4057)
/note="T7 promoter"
/note="promoter for bacteriophage T7 RNA polymerase"
promoter 4070..5281
/note="UbC promoter"
/note="human ubiquitin C promoter"
protein_bind 5294..5327
/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..6041
/codon_start=1
/product="reverse tetracycline transactivator 3"
/note="rtTA3"
/note="activates expression from Tet-responsive promoters
in the presence of doxycycline"
/translation="MSRLDKSKVINGALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
KNKRALLDALPIEMLDRHHTHFCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
DSMPPLLRQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
DALDDFDLDMLPG"
protein_bind 6042..6075
/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."
misc_feature 6098..6670
/note="IRES"
/note="internal ribosome entry site (IRES) of the
encephalomyocarditis virus (EMCV)"
CDS 6673..7275
/codon_start=1
/gene="pac from Streptomyces alboniger"
/product="puromycin N-acetyltransferase"
/note="PuroR"
/note="confers resistance to puromycin"
/translation="MATEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIE
RVTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGSRMAELSGSRL
AAQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLET
SAPRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
misc_feature 7291..7879
/note="WPRE"
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
CDS complement(7762..7773)
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/note="Factor Xa site"
/translation="IEGR"
LTR 8086..8319
/note="3 LTR (Delta-U3)"
/note="self-inactivating 3 long terminal repeat (LTR) from
HIV-1"
polyA_signal 8426..8650
/note="bGH poly(A) signal"
/note="bovine growth hormone polyadenylation signal"
rep_origin 8696..9124
/direction=RIGHT
/note="f1 ori"
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 9138..9467
/note="SV40 promoter"
/note="SV40 enhancer and early promoter"
rep_origin 9318..9453
/note="SV40 ori"
/note="SV40 origin of replication"
polyA_signal 10675..10796
/note="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
primer_bind complement(10845..10861)
/note="M13 rev"
/note="common sequencing primer, one of multiple similar
variants"
protein_bind 10869..10885
/bound_moiety="lac repressor encoded by lacI"
/note="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)."
promoter complement(10893..10923)
/note="lac promoter"
/note="promoter for the E. coli lac operon"
protein_bind 10938..10959
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding site"
/note="CAP binding activates transcription in the presence
of cAMP."
rep_origin complement(11247..11835)
/direction=LEFT
/note="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(12006..12866)
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/note="AmpR"
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
promoter complement(12867..12971)
/gene="bla"
/note="AmpR promoter"
polyA_signal 13019..13153
/note="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
