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RNAi Machinery Antibody Sampler Kit #4487

Kit Includes	Quantity	Applications	Reactivity	MW (kDa)	Isotype
Drosha (D28B1) Rabbit mAb #3364	40 µl	W IP	H M	160	Rabbit IgG
Argonaute 2 (C34C6) Rabbit mAb #2897	40 µl	W IP	H M R Mk	97	Rabbit IgG
Argonaute 1 (D84G10) XP® Rabbit mAb #5053	40 µl	W IP IHC-P	H M R Mk	97	Rabbit IgG
Mili (D14F5) XP® Rabbit mAb #5940	40 µl	W IP IHC-P IF-F	M	110	Rabbit IgG
Dicer (D38E7) Rabbit mAb #5362	40 µl	W IP	H	220	Rabbit IgG
Anti-rabbit IgG, HRP-linked Antibody #7074	100 µl				Goat

Applications Key:  W=Western Blotting  IP=Immunoprecipitation  IHC-P=Immunohistochemistry (Paraffin)  IF-F=Immunofluorescence (Frozen)
Reactivity Key:  H=Human  M=Mouse  R=Rat  Mk=Monkey
Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Specificity / Sensitivity

Each antibody in the RNAi Machinery Antibody Sampler Kit recognizes endogenous levels of their target protein and does not cross-react with other proteins.

Description

The RNAi Machinery Antibody Sampler Kit provides an economical means to analyze proteins associated with endogenous RNA interference. The kit contains enough primary and secondary antibodies to perform four western blot experiments.

Source / Purification

Polyclonal antibodies are produced by immunizing animals with synthetic peptides corresponding to the residues surrounding Ala1123 of human Dicer or residues surrounding Leu118 of mouse Mili protein. Polyclonal antibodies are purified by protein A and peptide affinity chromatography. Monoclonal antibody is produced by immunizing animals with synthetic peptides corresponding to the sequence of mouse Argonaute 2 or around Gly953 of human Drosha proteins.

Background

RNA interference (RNAi) serves as a global mechanism of gene regulation in eukaryotes. Through interactions with Dicer, Drosha, Argonaute 2 (Ago2) and Miwi/Mili proteins, microRNA (miRNA) is processed within the nucleus and utilized for gene silencing and down regulation of gene expression. Dicer is a member of the RNase III family that specifically cleaves double-stranded RNA to generate microRNA (miRNA) (1). Long, primary transcripts (pri-miRNAs) are processed to stem-looped pre-miRNAs by the nuclear RNase III Drosha (2) and are then transported to the cytoplasm for further processing by Dicer to produce mature, 22-nucleotide miRNAs (3). The mature miRNA then becomes a part of the RNA-Induced Silencing Complex (RISC) and can bind to the 3' UTR of the target mRNA (3). Interference of Drosha pri-miRNA processing results in the increase of pri-miRNAs and the decrease of pre-miRNAs (2). Drosha forms part of a multiprotein complex called the Microprocessor along with other components, such as DGCR8 (4). Both Drosha and DGCR8 are necessary for miRNA biogenesis (5). Argonaute protein family members participate in various steps of miRNA-mediated gene silencing such as repression of translation and mRNA turnover (6). The Drosophila piwi gene was identified as being required for the self-renewal of germ-line stem cells, and its homologues are well conserved among various species including Arabidopsis, C. elegans and human (7). Miwi and Mili proteins are both mouse homologs of Piwi and contain a carboxy-terminal Piwi domain that binds to Piwi-interacting RNAs (piRNAs) in male germ cells and are essential for spermatogenesis in mouse (8-11).

1. Hutvágner, G. and Zamore, P.D. (2002) Science 297, 2056-60.
2. Lee, Y. et al. (2003) Nature 425, 415-9.
3. Diederichs, S. and Haber, D.A. (2007) Cell 131, 1097-108.
4. Denli, A.M. et al. (2004) Nature 432, 231-5.
5. Gregory, R.I. et al. (2004) Nature 432, 235-40.
6. Peters, L. and Meister, G. (2007) Mol Cell 26, 611-23.
7. Cox, D.N. et al. (1998) Genes Dev 12, 3715-27.
8. Kuramochi-Miyagawa, S. et al. (2001) Mech Dev 108, 121-33.
9. Aravin, A. et al. (2006) Nature 442, 203-7.
10. Grivna, S.T. et al. (2006) Proc Natl Acad Sci U S A 103, 13415-20.
11. Grivna, S.T. et al. (2006) Genes Dev 20, 1709-14.

Application References

Have you published research involving the use of our products? If so we'd love to hear about it. Please let us know!

Protocols

2897:

Immunoprecipitation, Western Blotting

3364:

Immunoprecipitation, Western Blotting

5053:

IHC / Paraffin, Immunoprecipitation, Western Blotting

5362:

Immunoprecipitation, Western Blotting

5940:

IHC / Paraffin, Immunofluorescence, Immunoprecipitation, Western Blotting

Selected rabbit monoclonal antibodies are produced under license (granting certain rights including those under U. S. Patent No. 5,675,063) from Epitomics, Inc.

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For Research Use Only. Not For Use In Diagnostic Procedures.