BioProtocols

Telomere and telomerase assay

Saturday, 02 January 2010 06:26 biomed

A telomere is a region of repetitive DNA at the end of a chromosome, which protects the end of the chromosome from deterioration. Its name is derived from the Greek nouns telos (τἐλος) "end" and merοs (μέρος, root: μερ-) "part".

Russian theorist Alexei Olovnikov was the first to recognize (1971) the problem of how chromosomes could replicate right to the tip, as such was impossible with replication in a 5' to 3' direction. To solve this and to accommodate Leonard Hayflick's idea of limited somatic cell division, Olovnikov suggested that DNA sequences would be lost in every replicative phase until they reached a critical level, at which point cell division would stop.

Enucleation and Fusion Protocol for the Transfer of Mitochondrial DNA to rho-zero Cells

Saturday, 02 January 2010 05:46 ER

Once week prior to fusion start Rhodamine-6-G treatment of muscle. At the same time plate 2x10^5 of donor cells (5 plates).

1. The day before the actual fusion prepare the following Ficoll gradient (containing cytochalasin D or B for enucleation)

Ficall Final conc (%) 50% Ficoll stock 4xDMEM dH2O Cyto D (100ug/ml)

Protein Purification Protocols

Sunday, 20 December 2009 05:29 biomed

Protein purification is a series of processes intended to isolate a single type of protein from a complex mixture. Protein purification is vital for the characterisation of the function, structure and interactions of the protein of interest. The starting material is usually a biological tissue or a microbial culture. The various steps in the purification process may free the protein from a matrix that confines it, separate the protein and non-protein parts of the mixture, and finally separate the desired protein from all other proteins. Separation of one protein from all others is typically the most laborious aspect of protein purification. Separation steps exploit differences in protein size, physico-chemical properties and binding affinity.