y full sib families selected from the 200 broodstock families of

y full sib families selected from the 200 broodstock families of the Landcatch Natural Selection Atlantic salmon breeding program were specifically selected for the feeding trial. On the basis of parental genetic evaluations, 25 high flesh lipid contrasting with 25 low flesh lipid families were identified, and 35 fish from each family were transferred and grown in communal Trichostatin A sea water pens. All fish were tagged with electronic transponders to allow family identification while rearing in a common environment. After acclimation, the fish were grown for 12 weeks on the same low FM high VO diet containing 25% FM and 44% plant meals and a VO blend including rapeseed oil palm oil camelina oil. At the end of the trial, flesh samples were collected, frozen on dry ice and stored at ?20 C until lipid analysis.

Liver samples were also taken and stored at ?70 C for subsequent molecular analyses. Lipid analysis and choice of families for transcriptomic comparisons The 50 selected families were screened for their ability to retain and or synthesize n 3 LC PUFA when fed a low FM high VO diet. De boned and skinned flesh samples were combined into 3 pools per family for lipid analysis. Total lipids were extracted and determined gravimetrically from 1 2 g of pooled flesh. Fatty acid methyl esters were prepared by acid catalyzed transesterification of total lipids. Following purification, FAME were separated and quantified by gas liquid chromatography as described in. These data were used to select four families for transcriptomic analysis, two with equivalent high levels of lipid H, and two with equivalent low levels of lipid L.

Within each level of total lipid, two families with significantly con trasting relative n 3 LC PUFA levels were identified. RNA extraction and purification Hepatic tissue from ten individuals per family was rapidly homogenized in 2 ml TRI Reagent. Total RNA was isolated, following manufacturers instructions, and RNA quality and quantity was assessed by gel electro phoresis and spectrophotometry, respectively. Equal amounts of total RNA were pooled from two individuals to produce five biological replicates per family, which were further purified by mini spin column purification. Microarray hybridization and analysis A custom made Atlantic salmon oligoarray with 44 K features Drug_discovery per array on a four array per slide format, with experimental features printed singly was used.

The probes were co designed at the Institute of Aquaculture, University of Stirling, U. K. and Nofima, Norway, with array design available in the EBI Array Express database under accession number A MEXP 2065. The features were mainly derived from a core set of Atlantic salmon Unigenes supplemented with other unique cDNAs derived http://www.selleckchem.com/products/Perifosine.html from Genbank and the At lantic Salmon Gene Index. Probe annotations were derived from Blastx comparisons across four protein databases, as detailed elsewhere. The entire experiment com prised 20 hybridizations, 4 groups �� 5 biological replicates. Indire

Leave a Reply

Your email address will not be published. Required fields are marked *


You may use these HTML tags and attributes: <a href="" title=""> <abbr title=""> <acronym title=""> <b> <blockquote cite=""> <cite> <code> <del datetime=""> <em> <i> <q cite=""> <strike> <strong>