Hey
i am trying to isolate protein from wild type larvae and mutant to do some western blot , after 2 minutes the mutant extract starts turning black and within few minutes it turns completely black . Protein from wild type flies looks ok , i am not sure whats the reason , what i found on internet is that it could be because of phenol oxidase . but i want to know if someone has faced the same problem if yes ,how can i avoid blackening of the protein extract.
Thanks
Hello Khan, what method are you using for extraction? This could guide the answers you'll receive.
Meanwhile, you can follow these threads:
1. https://www.researchgate.net/post/What_is_the_protocol_for_protein_isolation_from_Drosophila_MelanogasterIsolation_of_protein_from_flies
2. https://www.google.be/url?sa=t&rct=j&q=&esrc=s&source=web&cd=5&cad=rja&uact=8&ved=0ahUKEwi08Pv39_HVAhXCYVAKHavCBEYQFghKMAQ&url=https%3A%2F%2Fwww.researchgate.net%2Fpost%2FCan_anyone_recommend_a_procedure_to_extract_proteins_from_adult_drosophila_heads&usg=AFQjCNGC7MfiT_YD1E7At-QbnhJZFbzHsg
3. https://www.google.be/url?sa=t&rct=j&q=&esrc=s&source=web&cd=6&cad=rja&uact=8&ved=0ahUKEwi08Pv39_HVAhXCYVAKHavCBEYQFghQMAU&url=https%3A%2F%2Fwww.researchgate.net%2Fpost%2FCan_anyone_suggest_a_method_to_extract_protein_from_drosophila_tissue&usg=AFQjCNEFRdHTJXaHxyS6MjSEto5IVmcRhw
Dear Robert
Thanks for the response , i am using lysis buffer ( 20mM Tris pH 7.6 , 150mM NaCl ,2mM EDTA, 10% glycerol , 1% Triton X- 100 , 1mM DTT and Protease inhibitor from Roche .
This was working fine for me but the mutant i am using now is showing this strange behavior . the wild type looks ok to me .
we still don't know the gene , we are trying to map the gene . its a new gene from our P-Element excision .
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