BI 117 Recitation Session 1. Techniques in Developmental Biology. Part 1 : Methods Using DNA, RNA, and Protein Jon or jev Kerckhoff 017 [email protected] The life codes. Methods with DNA and RNA. DNA->RNA->Protein - the flow of biological information. DNA = A, T, C, G. D: deoxyribose.
BI 117Recitation Session 1
Techniques in Developmental Biology
Part 1: Methods Using DNA, RNA, and Protein
Jon or jev
The life codes
Coding sequence of SpBra
Non-coding sequences (intergenic and intronic) in the vicinity of SpBra
Regulatory element binding sites
Purpose: to generate a superabundance of copies of your DNA fragment
Cloning is useful for many downstream applications!
Gene expression analysis
labeled dUTP (digoxigenin, biotin, fluorescein, etc.)
Substrate for Alkaline phosphatase
Methods with Proteins
Often conjugated to a flourescent molecule
Example of an immunohistochemistry figure
Embryos were stained at room temperature with the following primary antibodies: mouse monoclonal anti-Ftz, mouse monoclonal anti-Engrailed (a gift of N. H. Patel), and rabbit polyclonal anti-Even-skipped (a gift of M. Frasch). The primaries were visualized with Cy3 anti-mouse and FITC anti-rabbit
Genetics. 2004 September; 168(1): 161–180.
Immunoprecipitation (IP)The technique of precipitating a protein antigen out of solution using an antibody that specifically binds to that particular protein.Purpose: to isolate a specific protein from a lysate or crude extract
Histone H1 accumulation in sktl salivary glands. Salivary glands from wild-type (wt) and sktldrosophila larvae were dissected, analyzed by SDS-PAGE, and immunoblotted with histone H1 and tubulin antibodies. (Top) Antihistone H1 immunoblot. Histone H1 antibody recognizes both the phosphorylated (32 kD) and nonphosphorylated (31 kD) forms of histone H1. Nonphosphorylated histone H1 is completely absent in sktl mutant salivary glands. (Bottom) Same blot probed with tubulin antibody as a loading control.
From: Genetics, Vol. 167, 1213-1223, July 2004
Movie-The inner life in a cell
Embryology 101: An introduction to physical manipulations
Kuroda et al., 2005
Single Cell Labeling
Nature Reviews Genetics