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VPS39 Shrna Plasmid (H): Sc-90114-SH SANTA CRUZ BIOTECHNOLOGY, INC. VPS39 shRNA Plasmid (h): sc-90114-SH BACKGROUND STORAGE AND RESUSPENSION Vacuolar sorting proteins (VPSs) are required for proper trafficking of endocytic Store lyophilized shRNA plasmid DNA at 4° C with desiccant. Stable for and biosynthetic proteins to the vacuole and play an important role in the at least one year from the date of shipment. Once resuspended, store at budding process of cells. VPS39 (vacuolar protein sorting 39), also known as 4° C for short term storage or -80° C for long term storage. Avoid repeated VAM6 or TLP, is an 886 amino acid protein that localizes to the cytoplasm, as freeze thaw cycles. well as to the lysosomal and endosomal membrane, and contains one CNH Resuspend lyophilized shRNA plasmid DNA in 200 µl of the deionized water domain. Expressed ubiquitously with highest expression in kidney, heart, lung, provided. Resuspension of the shRNA plasmid DNA in 200 µl of deionized brain, placenta and skeletal muscle, VPS39 functions as a homooligomer that water makes a 0.1 µg/µl solution in a 10 mM Tris, 1 mM EDTA buffered is thought to play a role in the clustering and fusion of endosomes and lyso- solution. somes. Multiple isoforms of VPS39 exist due to alternative splicing events. The gene encoding VPS39 maps to human chromosome 15, which houses over APPLICATIONS 700 genes and comprises nearly 3% of the human genome. VPS39 shRNA Plasmid (h) is recommended for the inhibition of VPS39 REFERENCES expression in human cells. 1. Chapman, R.E. 1994. Vacuolar sorting. Tracking down an elusive receptor. SUPPORT REAGENTS Curr. Biol. 4: 1019-1022. For optimal shRNA Plasmid transfection efficiency, Santa Cruz Biotechnology’s 2. Nagase, T., Ishikawa, K., Suyama, M., Kikuno, R., Miyajima, N., Tanaka, shRNA Plasmid Transfection Reagent: sc-108061 (0.2 ml) and shRNA Plasmid A., Kotani, H., Nomura, N. and Ohara, O. 1998. Prediction of the coding Transfection Medium: sc-108062 (20 ml) are recommended. Control shRNAs sequences of unidentified human genes. XI. The complete sequences of are available as 20 µg lyophilized plasmid DNA. Each encodes a scrambled 100 new cDNA clones from brain which code for large proteins in vitro. shRNA sequence that will not lead to the specific degradation of any known DNA Res. 5: 277-286. cellular mRNA. Control shRNA Plasmids include: sc-108060, sc-108065 and 3. Hartley, J.L., Temple, G.F. and Brasch, M.A. 2000. DNA cloning using in sc-108066. vitro site-specific recombination. Genome Res. 10: 1788-1795. GENE EXPRESSION MONITORING 4. Caplan, S., Hartnell, L.M., Aguilar, R.C., Naslavsky, N. and Bonifacino, J.S. 2001. Human VAM6P promotes lysosome clustering and fusion in vivo. J. VPS39 (C-5): sc-514762 is recommended as a control antibody for monitoring Cell Biol. 154: 109-122. of VPS39 gene expression knockdown by Western Blotting (starting dilution 1:200, dilution range 1:100-1:1000) or immunofluorescence (starting dilution CHROMOSOMAL LOCATION 1:50, dilution range 1:50-1:500). Genetic locus: VPS39 (human) mapping to 15q15.1. To ensure optimal results, the following support reagents are recommended: 1) Western Blotting: use m-IgGκ BP-HRP: sc-516102 or m-IgGκ BP-HRP (Cruz PRODUCT Marker): sc-516102-CM (dilution range: 1:1000-1:10000), Cruz Marker™ Molecular Weight Standards: sc-2035, UltraCruz® Blocking Reagent: VPS39 shRNA Plasmid (h) is a pool of 3 target-specific lentiviral vector plas- sc-516214 and Western Blotting Luminol Reagent: sc-2048. 2) Immunofluo- mids each encoding 19-25 nt (plus hairpin) shRNAs designed to knock down rescence: use m-IgGκ BP-FITC: sc-516140 or m-IgGκ BP-PE: sc-516141 gene expression. Each vial contains 20 µg of lyophilized shRNA plasmid DNA. (dilution range: 1:50-1:200) with UltraCruz® Mounting Medium: sc-24941 Suitable for up to 20 transfections. Also see VPS39 siRNA (h): sc-90114 and or UltraCruz® Hard-set Mounting Medium: sc-359850. VPS39 shRNA (h) Lentiviral Particles: sc-90114-V as alternate gene silencing products. RT-PCR REAGENTS RESEARCH USE Semi-quantitative RT-PCR may be performed to monitor VPS39 gene expres- sion knockdown using RT-PCR Primer: VPS39 (h)-PR: sc-90114-PR (20 µl). The purchase of this product conveys to the buyer the nontransferable right Annealing temperature for the primers should be 55-60° C and the exten- to use the purchased amount of the product and all replicates and derivatives sion temperature should be 68-72° C. for research purposes conducted by the buyer in his laboratory only (whether the buyer is an academic or for-profit entity). The buyer cannot sell or other- wise transfer (a) this product (b) its components or (c) materials made using PROTOCOLS this product or its components to a third party, or otherwise use this product See our web site at www.scbt.com for detailed protocols and support or its components or materials made using this product or its components products. for Commercial Purposes. Santa Cruz Biotechnology, Inc. 1.800.457.3801 831.457.3800 fax 831.457.3801 Europe +00800 4573 8000 49 6221 4503 0 www.scbt.com.
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