JSU Student Symposium 2020
 
Blocking Buffers and Their Effects on Mammalian Histones

Date

2-13-2020

Faculty Mentor

Roger Sauterer, Biology

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Submission Type

Poster

Location

Houston Cole Library, 11th Floor | 4:00-4:10 p.m.

Description

Mammalian histones are highly alkaline proteins found in the nuclei that organize DNA into chromosomes and regulate transcription. We are investigating the interactions between histones and mitochondria by using cell fractionation and Western Blotting to identify the histones. The blocking step in the process coats the membrane with proteins or other molecules to reduce non-specific binding of the antibodies. We use 5% nonfat dry milk as a control for all experiments, it is essential to use this variation of milk because it reduces background noise and helps produce good, clear bands. Although 5% nonfat dry milk is widely used as a blocking buffer, we are testing different blocking buffers to see if the signal strength increases or completely strips proteins off of the blot, the conclusion of which blocking buffer works the best will be determined by the protein being examined. Since antibodies to histone H2A and H2B are low-affinity and give a weak signal, we are therefore testing different blocking buffers, such as milk concentrations ranging from 1 to 5%, BSA ranging from 1 to 5%, and hemoglobin ranging from 1 to 5%, as well as gelatin at 3%, and PVP or PEG individually ranging from 1 to 4% and different combinations of the aforementioned blocking buffers to determine which blocking buffer gives the best detection signal. Our preliminary results indicated that RVR/REG combinations provided the strongest signal, while gelatin tends to strip the proteins completely off of the blot and reduce the signal.

Keywords

student presentations, student posters, histones, blocking buffers

Rights

This video is the property of Jacksonville State University and is intended for non-commercial use. Video and images may be copied for personal use, research, teaching or any "fair use" as defined by copyright law. Users are asked to acknowledge Jacksonville State University. For more information, please contact digitalcommons@jsu.edu.

Disciplines

Biology

Presentation Information

Tavis, P. and Harris, S. (2020, 13 February). Blocking Buffers and their effects on mammalian histones. Poster presented at the 2020 JSU Student Symposium, Jacksonville State University, Jacksonville, AL.

Blocking Buffers and Their Effects on Mammalian Histones

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