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VPS39 Shrna (H) Lentiviral Particles: Sc-90114-V SANTA CRUZ BIOTECHNOLOGY, INC. VPS39 shRNA (h) Lentiviral Particles: sc-90114-V BACKGROUND APPLICATIONS Vacuolar sorting proteins (VPSs) are required for proper trafficking of endocytic VPS39 shRNA (h) Lentiviral Particles is recommended for the inhibition of and biosynthetic proteins to the vacuole and play an important role in the VPS39 expression in human cells. budding process of cells. VPS39 (vacuolar protein sorting 39), also known as VAM6 or TLP, is an 886 amino acid protein that localizes to the cytoplasm, as SUPPORT REAGENTS well as to the lysosomal and endosomal membrane, and contains one CNH Control shRNA Lentiviral Particles: sc-108080. Available as 200 µl frozen domain. Expressed ubiquitously with highest expression in kidney, heart, lung, viral stock containing 1.0 x 106 infectious units of virus (IFU); contains an brain, placenta and skeletal muscle, VPS39 functions as a homooligomer that shRNA construct encoding a scrambled sequence that will not lead to the is thought to play a role in the clustering and fusion of endosomes and lyso- specific degradation of any known cellular mRNA. somes. Multiple isoforms of VPS39 exist due to alternative splicing events. The gene encoding VPS39 maps to human chromosome 15, which houses over GENE EXPRESSION MONITORING 700 genes and comprises nearly 3% of the human genome. VPS39 (C-5): sc-514762 is recommended as a control antibody for monitoring REFERENCES of VPS39 gene expression knockdown by Western Blotting (starting dilution 1:200, dilution range 1:100-1:1000) or immunofluorescence (starting dilution 1. Chapman, R.E. 1994. Vacuolar sorting. Tracking down an elusive receptor. 1:50, dilution range 1:50-1:500). Curr. Biol. 4: 1019-1022. To ensure optimal results, the following support reagents are recommended: 2. Nagase, T., Ishikawa, K., Suyama, M., Kikuno, R., Miyajima, N., Tanaka, 1) Western Blotting: use m-IgGκ BP-HRP: sc-516102 or m-IgGκ BP-HRP (Cruz A., Kotani, H., Nomura, N. and Ohara, O. 1998. Prediction of the coding Marker): sc-516102-CM (dilution range: 1:1000-1:10000), Cruz Marker™ sequences of unidentified human genes. XI. The complete sequences of Molecular Weight Standards: sc-2035, UltraCruz® Blocking Reagent: 100 new cDNA clones from brain which code for large proteins in vitro. sc-516214 and Western Blotting Luminol Reagent: sc-2048. 2) Immunofluo- DNA Res. 5: 277-286. rescence: use m-IgGκ BP-FITC: sc-516140 or m-IgGκ BP-PE: sc-516141 3. Hartley, J.L., Temple, G.F. and Brasch, M.A. 2000. DNA cloning using in (dilution range: 1:50-1:200) with UltraCruz® Mounting Medium: sc-24941 vitro site-specific recombination. Genome Res. 10: 1788-1795. or UltraCruz® Hard-set Mounting Medium: sc-359850. 4. Caplan, S., Hartnell, L.M., Aguilar, R.C., Naslavsky, N. and Bonifacino, J.S. RT-PCR REAGENTS 2001. Human VAM6P promotes lysosome clustering and fusion in vivo. J. Cell Biol. 154: 109-122. 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). 5. Felici, A., Wurthner, J.U., Parks, W.T., Giam, L.R., Reiss, M., Karpova, T.S., Annealing temperature for the primers should be 55-60° C and the exten- β McNally, J.G. and Roberts, A.B. 2003. TLP, a novel modulator of TGF sig- sion temperature should be 68-72° C. naling, has opposite effects on Smad2- and Smad3-dependent signaling. EMBO J. 22: 4465-4477. BIOSAFETY 6. Online Mendelian Inheritance in Man, OMIM™. 2008. Johns Hopkins Lentiviral particles can be employed in standard Biosafety Level 2 tissue University, Baltimore, MD. MIM Number: 612188. World Wide Web URL: culture facilities (and should be treated with the same level of caution as http://www.ncbi.nlm.nih.gov/omim/ with any other potentially infectious reagent). Lentiviral particles are repli- cation-incompetent and are designed to self-inactivate after transduction CHROMOSOMAL LOCATION and integration of shRNA constructs into genomic DNA of target cells. Genetic locus: VPS39 (human) mapping to 15q15.1. RESEARCH USE PRODUCT The purchase of this product conveys to the buyer the nontransferable right VPS39 shRNA (h) Lentiviral Particles is a pool of concentrated, transduction- to use the purchased amount of the product and all replicates and derivatives ready viral particles containing 3 target-specific constructs that encode for research purposes conducted by the buyer in his laboratory only (whether 19-25 nt (plus hairpin) shRNA designed to knock down gene expression. Each the buyer is an academic or for-profit entity). The buyer cannot sell or other- vial contains 200 µl frozen stock containing 1.0 x 106 infectious units of virus wise transfer (a) this product (b) its components or (c) materials made using (IFU) in Dulbecco’s Modified Eagle’s Medium with 25 mM HEPES pH 7.3. this product or its components to a third party, or otherwise use this product Suitable for 10-20 transductions. Also see VPS39 siRNA (h): sc-90114 and or its components or materials made using this product or its components VPS39 shRNA Plasmid (h): sc-90114-SH as alternate gene silencing products. for Commercial Purposes. STORAGE PROTOCOLS Store lentiviral particles at -80º C. Stable for at least one year from the See our web site at www.scbt.com for detailed protocols and support date of shipment. Once thawed, particles can be stored at 4º C for up to products. one week. Avoid repeated freeze thaw cycles. 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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