pSINsi-hH1 DNA
Contents
1. e DNA mesure v201104 TaKaRa 7s Tel 077 543 6116 bx 07784319792. 5 LTR mRNA polll RNA SV40 RNA Dicer siRNA pSINsi 3 pSINsi hH1 DNA 3660 human H1 promoter Accession S68670 pSINsi hU6 DNA 3661 humanU6 promoter Accession X07425 PSINsi mU6DNA 3662 mouse U6 promoter Accession X06980 0 _ 10mM Tris HCl pH8 0 1mM EDTA E 6355bp 1 dideoxy human H1 promoter Accession S68670 2 gz77H 1 1 BamH1 Clal S
3. Lot No pSINsi RNA polymeraselll poll RNA pollll RNA DNA siRNA siRNA Retrovirus Packaging Kit Eco Ampho 6160 6161 pSINsi 3 LTR U3
4. Ori 6 355 bp stuffer coR Bin In Nco Ao al MLV US a SM40 MLVR R N romoter ee U3 eo p 3 LTR NOTICE TO PURCHASER LIMITED LICENSE L21 pDON Vecter This product is covered by the claims of U S Patent No 6 190 907 6 451 595 and their foreign counterpart patent claims Note This product is for research use only It is not intended for use in therapeutic or diagnostic procedures for humans or animals Also do not use this product as food cosmetic or household item etc Takara products may not be resold or transferred modified for resale or transfer or used to manufacture commercial products without written approval from TAKARA BIO INC If you require licenses for other use please contact us by phone at 81 77 543 7247 or from our website at www takara bio com The viral supernatants produced by this retroviral vector could con tain potentially hazardous recombinant virus depending on an insert gene Due caution must be exercised in this production and handling of recombinant retrovirus Please follow the guideline for experiments using recombinant DNA issued by the relevant authorities and the safety committee of your organization or your country in using this product Takara is not liable for any accident or damage caused by the use of this product v201104 pSINsi hH1 DNA Code No 3660 Size 20 ug Shipping at 20 C Store at 20 C 2
5. h different promoter A customer can choose an appropriate vector depending on a kind of target cell and an experimental purpose pSINsi hH1 DNA Cat 3660 contains human H1 promoter Accesion S68670 pSINsi hU6 DNA Cat 3661 contains human U6 promoter Accession X07425 pSINsi mU6 DNA Cat 3222 contains mouse U6 promoter Accession X06980 Form 10mM Tris HCl pH8 0 1mM EDTA Preparation Purified by ion exchange column treatment Chain length 6 355 bp Purity Confirmed that the promoter on this vector is the same as human H1 promoter Accession S68670 by dideoxy sequencing method Confirmed to be cleaved at a single site by restriction enzymes BamHI Also confirmed that a Stuffer is excised with BamH I Cla Confirmed that the vector is transfected into 293T cell with Retrovirus Packaging Kit and then virus vector is emitted out of the cell Applications Retrovirus mediated siRNA expression in mammalian cells Reference 1 Brummelkamp et a 2002 Science 296 550 553 2 Lee et al 2002 Nat Biotech 20 500 505 3 Paddison et a 2002 Genes and Dev 16 948 958 4 Paul et a 2002 Nat Biotech 20 505 508 5 Sui et al 2002 Proc Natl Acad Sci USA 99 5515 5520 6 Kawasaki et a 2003 Nucleic Acids Res 31 700 707 Vector map 5 LTR HCMV IE promoter MLV R MLVU5 Hind Ill SD Amp Nco y EcoRI promoter Hind Ill ae aed H1 pSINsi hH1 ean
6. pSINsi hH1 DNA Code No 3660 Size 20 ug Shipping at 20 C Store at 20 C 2years from date of receipt under proper storage conditions Lot No Concentration 9 ul Volume yl Regarding protocol please refer to the product manual supplied with this vector Description pSINsi vector series is a vector for expression of hairpin RNA using a RNA polymerase III pol Ill promoter This vector series was developed from a self inactive retrovirus vector plasmid and contains neomycin resistant gene as a marker By inserting synthesized oligo DNA sequence for hairpin RNA expression into the downstream of the pol Ill promoter siRNA expres sion retrovirus vector plasmid is constructed The obtained retrovirus vec tor plasmid for siRNA expression is used to prepare recombinant retrovirus with Retrovirus Packaging Kit Eco Ampho Cat 6160 6161 The retrovirus vector plasmids of pSINsi vector series are deleted with the promoter activity of 3 LTR U3 region In the state of provirus incorporated into chromosome after transcribed 5 LTR promoter activity disappears and mRNA derived from provirus cannot be transcribed Accordingly only the hairpin RNA from the pol Ill promoter and the neomycin resistant gene from SV40 promoter are transcribed The expressed hairpin RNA is prepared to siRNA through digestion by a Dicer present in a cell Three kinds of plasmid vector are available in pSINsi vector series Each carries eac
7. tuffer 3 Retrovirus Packaging Kit 293T siRNA 1 Brummelkamp etal 2002 Science 296 550 553 2 Lee et a 2002 Nat Biotech 20 500 505 3 Paddison et a 2002 Genes and Dev 16 948 958 4 Paul et a 2002 Nat Biotech 20 505 508 5 Sui et a 2002 Proc Natl Acad Sci USA 99 5515 5520 6 Kawasaki et a 2003 Nucleic Acids Res 31 700 707 pSINsi hH1 DNA 5 LTR HCMV IE promoter MLV R MLV U5 Hind III SD Amp co Y EcoRI promoter Hind M 8 human H1 a pSINsi hH1 p anmi Ori 6 355 bp stuffer EcoRI Bin In MLV ae a oa a SV40 MLV R Neo promoter deleted U3 3 LTR
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