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QTL for CLA content in the milk in the Sarda x Lacaune population

Conjugated linoleic acid (CLA) is a collective term for positional and geometric isomers of linoleic acid (C18:2). The cis 9 trans 11 C18:2 (rumenic acid) is the most abundant CLA isomer in meat and milk of ruminants. A portion of CLA is formed in the rumen by biohydrogenation of linoleic acid, escapes further biohydrogenation and is absorbed in the digestive tract. The extent of this process is minimal, while the intermediate product of CLA biohydrogenation, (trans 11-C18:1; vaccenic acid), accumulates. The other important source of vaccenic acid (VA) in the rumen is the biohydrogenation of the linolenic acid (C18:3). This pathway does not have as intermediate CLA. VA produced in the rumen is desaturated to produce CLA in the mammary gland tissues by Stearoyl-CoA desaturase (Scd). Scd can use different fatty acids as substrate and influences the amount of several unsaturated fatty acids. On the whole, there is evidence that milk CLA basically comes from mammary synthesis by the action of Scd on VAAC provided by the rumen. Thus differences in ruminant species, cattle breeds and between individual cows can probably be explained by the different activities of Scd, even when they were fed with the same diet.

On the Sarda x Lacaune backcross ewes, individual samplings to determine FA content in milk were performed twice, in the middle of the 2nd and the 3rd lactations. Fatty acid content was determined as follows: fat separation by centrifuging at low temperature, storage of individual cream at 20°C, oil separation by thermal shock and centrifuging, acid trans-methylation. Fatty acid methyl esters (FAME) were determined by gas chromatography using a VARIAN GC 3600 equipped with FID and a fused silica capillary column (SP 2560 Supelco), 100 m  0.25 mm i.d., film thickness 0.20 µm. Helium was used as the carrier gas at a flow of 1 ml/min. The split ratio was 1:100. The oven temperature was programmed at 75°C and held for 1.50 min, then increased to 190°C at a rate of 8°C/min, held for 25 min, increased to 230°C at 15°C/min, held for 4.47 min. The temperatures of the injector and of the detector were set at 290°C. For QTL detection analyisis 2 traits were considered: the CLA content in the milk fat (mg/g of fat) and the ratio between CLA and vaccenic acid. The latter is the direct precursor of CLA in the mammary gland where it is desaturated by delta 9-desaturase to produce rumenic acid. A repeated measurement model including the fixed effect year x group of management and the random effects of individual and sire was applied prior to QTL analysis. An across family single trait QTL analysis was carried out, using the methodology proposed by Knott et al. (1996) and Elsen et al. (1999), by within-sire linear regression and implemented in the QTL map software.

The CLA content was consistent with previous results in Sarda sheep, although higher values of CLA (32.85 mg/g of fat) were found in Sarda sheep grazing during the spring. CLA content showed remarkable variability between families. Indeed sire variance was 8.4 % of the total phenotypic variance for the CLA content and 5.6 % for the ratio CLA/VACC. Total individual variance was 29.7 % for CLA content and 34.4 % for the ratio. Three QTL were detected for the CLA content in milk fat on OAR 4, 14, and 19. The effects ranged from 0.33 to 0.83, on average between 47% and 124% of the overall rsd. Only three families per QTL were significant. Two of the four QTL identified for CLA/VACC, on OAR 4 and 14, are in the same regions of those found for CLA. This suggests that these QTL affect CLA above all. By contrast, the QTL found on OAR 6 and 22 were specific to the ratio.

This indicates that CLA/VACC, which reflects the rate of desaturation of the vaccenic acid in rumenic acid, can be considered a specific trait. The number of significant families ranged from two to four with effects of around two thirds of the overall rsd. Of particular interest is the QTL detected on OAR 22 where the SCD gene encoding for delta 9-desaturase is located. This result can be considered as a first step, and suggests that SCD gene polymorphism is actually related to the delta 9-desaturase expression level and affects the quantity of CLA produced in the mammary tissues from vaccenic acid. The coincidence between the most probable location and the map position of the Scd gene supports future research aimed at finding causal mutations along that gene. Thus a fine mapping of OAR22 has been undertaken and the sequencing of the candidate gene has been planned for the next future.

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Ricercatore Genetica Applicata alle Produzioni Animali
Loc. Bonassai - S.S. Sassari-Fertilia Km 18,6
07040 Olmedo (SS)
Italy
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