Anti lexa

Manufactured by Takara Bio

Anti-LexA is a laboratory reagent used to detect and quantify the presence of the LexA protein. LexA is a transcriptional regulator involved in the SOS response, a cellular mechanism that responds to DNA damage. Anti-LexA can be used in various experimental techniques, such as Western blotting, immunoprecipitation, and enzyme-linked immunosorbent assays (ELISA), to study the dynamics and regulation of the LexA protein.

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4 protocols using anti lexa

Protein Extraction and Immunoblotting from Diverse Organisms

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Total protein extraction from yeast, E. coli, and rice plants and immunoblotting (Western blotting) were performed as previously described (Chen et al., 2010b (link)). The anti-OsSERK2 antibody against the synthetic peptide AELAPRHNDW-Cys of OsSERK2 (amino acids 602–611) was provided as a service by Pacific Immunology. Detailed information about their methods can be obtained at Pacific Immunology (www.pacificimmunology.com/). Anti-OsSERK2 for detection of OsSERK2, anti-LexA (Clontech) for detection of LexA-fused protein produced from BD vectors, anti-HA (Covance) for detection of HA-fused protein produced from AD vectors, and anti-Myc (Santa Cruze Biotech) for detection of XA21 with Myc tag were used as primary antibodies.

Chen X., Zuo S., Schwessinger B., Chern M., Canlas P.E., Ruan D., Zhou X., Wang J., Daudi A., Petzold C.J., Heazlewood J.L, & Ronald P.C. (2014). An XA21-Associated Kinase (OsSERK2) Regulates Immunity Mediated by the XA21 and XA3 Immune Receptors. Molecular Plant, 7(5), 874-892.

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Antibody Generation and Validation Protocols

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Rabbit anti-Robo1 and anti-Arl4A/C/D antibodies were generated as described previously (Li et al., 2007 (link)). The following antibodies were used: anti-Myc (1:1000, catalogue no. MMS-150R; Covance, Princeton, NJ), anti-HA (1:1000, catalogue no. SC-7392; Santa Cruz Biotechnology), anti-LexA (1:1000, catalogue no. 5397-1; Clontech, Mountain View, CA), anti-CD44 (1:200 for IF, catalogue no. MA4400; Invitrogen), anti-Cdc42 (1:500, catalogue no. 2462; Cell Signaling, Danvers, MA), anti-srGAP1 (1:1000, catalogue no. 76926; Abcam), and anti-α-tubulin (1:5000, catalogue no. T5168; Sigma-Aldrich, St. Louis, MO). A blue-fluorescent DNA stain 4′,6-diamidino-2-phenylindole (DAPI) solution was purchased from Millipore (1:5000, catalogue no. S7113). Horseradish peroxidase (HRP)–conjugated goat anti-rabbit and anti-mouse antibodies were purchased from GE Healthcare (1:5000, catalogue nos. NA934V and NA931V, respectively; Waukesha, WI). Alexa Fluor–conjugated secondary antibodies were purchased from Invitrogen (Grand Island, NY). Alexa Fluor 594/488–conjugated anti-rabbit/anti-mouse and Alexa Fluor 647–conjugated anti-rat secondary antibodies were from Invitrogen (1:1000, catalogue nos. A-11012 for Alexa Fluor 594-rabbit, A-11034 for Alexa Fluor 488-rabbit, A-11001 for Alexa Fluor 488-mouse, A-11032 for Alexa Fluor 594-mouse, and A-21247 for Alexa Fluor 647-rat).

Chiang T.S., Lin M.C., Tsai M.C., Chen C.H., Jang L.T, & Lee F.J. (2019). ADP-ribosylation factor–like 4A interacts with Robo1 to promote cell migration by regulating Cdc42 activation. Molecular Biology of the Cell, 30(1), 69-81.

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Antibody Protocol for Arl4A/C/D

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The preparation of polyclonal antibodies against Arl4A/C/D was described previously (Li et al., 2007 (link)). The following primary antibodies were used: anti-FLNa (1:1000, catalogue MAB1678; Millipore, Billerica, MA), anti-alpha tubulin (1:5000, catalogue T5168; Sigma, St. Louis, MO), anti-fascin (1:100, catalogue MAB3582; Millipore), anti-FGD6 (1:200, SC-167891; Santa Cruz Biotechnology, Santa Cruz, CA), anti-Rac1 (1:1000, catalogue 05-389; Millipore), anti-Cdc42 (1:250, catalogue 610929; BD Biosciences, San Jose, CA), anti-myc (1:1000, catalogue MMS-150R; Covance, Princeton, NJ), anti-HA (1:1000, catalogue SC-7392; Santa Cruz Biotechnology), anti-LexA (1:1000, catalogue 5397-1; Clontech, Mountain View, CA), and anti–phalloidin-594 and -488 (1:500, catalogue 8953 and 8878; Cell Signaling). 4’,6-Diamidino-2-phenylindole (DAPI) solution was purchased from Millipore (1:5000, catalogue S7113). Horseradish peroxidase–conjugated goat anti-rabbit and anti-mouse antibodies were purchased from GE Healthcare (1:5000, catalogue NA934V and NA931V; Waukesha, WI). Alexa Fluor 594– and 488–conjugated anti-rabbit or anti-mouse secondary antibodies were purchased from Invitrogen (1:500, catalogue A-11012 for Alexa Fluor 594-rabbit; A-11034 for Alexa Fluor 488-rabbit; A-11001 for Alexa Fluor 488-mouse; A-11032 for Alexa Fluor 594-mouse; Grand Island, NY)

Chiang T.S., Wu H.F, & Lee F.J. (2017). ADP-ribosylation factor–like 4C binding to filamin-A modulates filopodium formation and cell migration. Molecular Biology of the Cell, 28(22), 3013-3028.

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Western Blot Analysis of Protein Extracts

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Total protein extracts from yeast, Xoo and rice plants and Western blot analyses were performed as previously described [41 (link),54 (link)]. The primary antibodies used were as follows: Anti-OsSERK2 for detection of OsSERK2 [54 (link)], anti-GFP (Santa Cruze Biotech) for detection of EFR::GFP and EFR::XA21::GFP, anti-LexA (Clontech) for detection of LexA-fused proteins expressed in yeast from pLexA, anti-HA (Covance) for detection of HA-tagged proteins expressed in yeast from pB42AD and anti-EF-Tu (Thermo Fisher Scientific, PA5-27512) antibody to detect EF-Tu in Xoo protein preparations. The appropriate secondary antibody, anti-mouse (Santa Cruz Biotech) and anti-rabbit (GE Healthcare) coupled to horseradish peroxidase were used in combination with chemiluminescence substrates (Thermo) to detect proteins by exposure to film.

Schwessinger B., Bahar O., Thomas N., Holton N., Nekrasov V., Ruan D., Canlas P.E., Daudi A., Petzold C.J., Singan V.R., Kuo R., Chovatia M., Daum C., Heazlewood J.L., Zipfel C, & Ronald P.C. (2015). Transgenic Expression of the Dicotyledonous Pattern Recognition Receptor EFR in Rice Leads to Ligand-Dependent Activation of Defense Responses. PLoS Pathogens, 11(3), e1004809.

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