Antibody Generation and Characterization for PpV

The PpV coding sequence was cloned into an expression vector with a C-terminal His-tag. The PpV-His protein was purified under denaturing conditions (Trenzyme, Konstanz). Rabbits were immunized with the purified denatured protein (BioGenes, Berlin). In western blots the serum detected a band that was not present in extracts from PpV embryos. In whole mount staining no difference between wild type and PpV embryos was observed, indicating unspecific background staining. Following antibodies were used: AuroraA (Giet et al. 2002 (link)), Feo (rabbit) (Vernì et al. 2004 (link)), LaminDm0 (mouse, T47/1/1) (Risau et al. 1981 (link)), γ-Tubulin (GTU-88, Sigma-Aldrich), GFP-booster (Chromotek), α-Tubulin (mouse; Sigma-Aldrich), CID (rabbit) (Jäger et al. 2005 (link)), P-D-TACC (rabbit) (Barros et al. 2005 (link)), Eve (Guinea pig, immunization according to (Frasch and Levine 1987 (link))), pH 3 (mouse, Millpore) and Dia (rabbit, guinea pig) (Grosshans et al. 2005 (link); Wenzl et al. 2010 (link)) .

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Liu B., Sung H.W, & Großhans J. (2019). Multiple Functions of the Essential Gene PpV in Drosophila Early Development. G3: Genes|Genomes|Genetics, 9(11), 3583-3593.

Publication 2019

γ-Tubulin (GTU-88, GFP-booster α-Tubulin

Antibodies Booster Coding sequence Embryos Guinea pig Immunization Mouse Protein Rabbit Rabbits Serum Tubulin Vector Western blots α tubulin

Corresponding Organization : Philipps University of Marburg

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Variable analysis

independent variables

Purified denatured PpV-His protein used for immunization of rabbits

dependent variables

Detection of a band in western blots that was not present in extracts from PpV embryos
Difference in whole mount staining between wild type and PpV embryos

control variables

Extracts from PpV embryos as a negative control for western blot
Wild type embryos as a control for whole mount staining

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