Optimised technique aids research in tough tissues
A protocol to extract genetic material from hard or frozen tissue will aid research in a range of species.
A research team has developed a protocol to study the genetic makeup of tough tissue, such as fish skin and fins.
Their technique, which optimises an existing method of extracting the nucleus from cells to study their DNA and RNA, meets a need for a standardised practice to remove the nucleus from a variety of tissue cells. This allows scientists to study the cells’ genome, helping them understand how cells function.
This method was developed to work with Atlantic salmon, as obtaining good quality nuclei from skin samples has previously been challenging, experts report.
The protocol, from a Roslin Institute team, has since been adapted for use in different tough or flash-frozen tissue and in a variety of species, such as mice, sharks, rabbits, shrimp and sea lice.
Scientists sought to develop a method that enables sample quality to be preserved, without using enzymes or heat which can damage cells, especially in cold-blooded species such as fish.
This method isolates the cell nuclei by putting frozen and minced tissue samples in a specialised salt solution, and filtering the solution until debris are removed and only the nuclei remain.
Specialised equipment then reads genetic information from the individual nuclei, facilitating the preparation of libraries for genomic sequencing.
Roslin experts identified the need for a protocol to apply to tissue such as Atlantic salmon skin, which is difficult to work with due to its toughness, the presence of connective tissue and fat deposits.
This method has been made widely accessible to the scientific community through a step-by-step guide, which researchers around the world have applied to a range of tissue types.
This research was published in PLOS One.
This protocol was developed to allow us to understand what cells are doing in their natural state, without being disturbed by removing them from tissues.
"We use this method here in Roslin, and we've had collaborators from across the world telling us they use it. It’s given us a wide network of collaborators and researchers that are taking up this methodology.
This method is broadly applicable to most tissue types, improving previous protocols in terms of consistency, cost-efficiency and nuclei quality.
"This optimised method has allowed us to study sea lice resistance across four salmonid species using a technology known as single-nuclei RNA sequencing, with potential massive benefits for fish welfare.
** The Roslin Institute receives strategic investment funding from the Biotechnology and Biological Sciences Research Council and it is part of the University of Edinburgh’s Royal (Dick) School of Veterinary Studies. **