Potential utility of 14 candidate housekeeping genes as normalization reference for RT-qPCR analysis with developmental samples (fertilized eggs to late veliger larvae) in Pacific abalone Haliotis discus hannai was evaluated using four different statistical algorithms (geNorm, NormFinder, BestKeeper and comparative ΔCT method). Different algorithms identified different genes as the best candidates, and geometric mean-based final ranking from the most to the least stable expression was as follow: RPL5, RPL4, RPS18, RPL8, RPL7, UBE2, RPL7A, GAPDH, RPL36, PPIB, EF1A, ACTB and B-TU. The findings were further validated via relative quantification of metallothionein (MT) transcripts using the stable and unstable reference genes, and expression levels of MT were greatly influenced according to the choice of reference genes. In overall, our data suggest that RPL5 and RPS18, either singly or in combination, are appropriate for normalizing gene expression in developmental samples of this abalone species, whereas ACTB, B-TU and EF1A are less stable and not recommended. In addition, our findings propose that standard deviations in geometric ranking as well as geometric mean itself should also be taken into account for the final selection of reference gene(s). This study could be a useful basis to facilitate the generation of accurate and reliable RT-qPCR data with developmental samples in this abalone species.

To investigate the effect of carnosine on exhaustive exercise, swimming tests were conducted weekly with loads corresponding to 5% of body weight attached to the tails of mice, and the swimming time to exhaustion was measured. Eighty male ICR mice were divided into four groups, to which carnosine was administered at doses of 0 (control), 10, 50, and 250 mg/kg/day, respectively, for a period of four weeks. At the end of swimming exercise challenges, serum biochemistry, oxidative stress enzyme activity, and antioxidant enzyme activity in tissues were determined. Treatment with 250 mg/kg carnosine resulted in a significant increase in swimming times to exhaustion, compared to the control group in the first (P<0.01) and third week (P<0.05). Significantly lower serum lactate levels were observed after the swimming exercise in the carnosine-treated groups (10 and 250 mg/kg), compared with the control (P<0.01). Malondialdehyde levels in the liver (10 and 50 mg/kg carnosine treated groups) and skeletal muscle (250 mg/kg carnosine treated group) were significantly lower, compared with the control (P<0.05). Significantly lower protein carbonyl levels in skeletal muscle were observed in the 50 and 250 mg/kg carnosine treated groups, compared with the control (P<0.01). Superoxide dismutase and glutathione peroxidase activities in skeletal muscle did not differ significantly among the groups. These results indicate that carnosine may improve swimming exercise capacity by attenuating production of lactate and reducing oxidative stress in mice.

(E,E)-8,10-dodecadienyl acetate (E8E10-12:Ac) and (E)-8-dodecaenyl acetate (E8-12:Ac) have been selected as the candidate chemicals for sex pheromone components of the M. phaseoli, female through GC-EAD tests, whereas the two compounds and an additional candidate, (E,Z)-7,9-dodecadienyl acetate (E7Z9-12:Ac), have been found at a ratio of 7:1:1 in the abdominal tip extract (Yum et al., 2008). In order to determine the actual composition of sex pheromone, therefore, several blends using the three chemicals were evaluated for attractiveness to males of M. phaseoli around red bean and soybean fields. Individual components as well as two blends consisted of E8E10-12:Ac/E7Z9-12:Ac and E8-12:Ac/E7Z9-12:Ac did not show attractiveness, whereas the blend of E8E10-12:Ac/E8-12:Ac showed an increased effect in male capture. Of the tested blends with all three chemicals, the 7:1:2 composition of E8E10-12:Ac, E8-12:Ac and E7Z9-12:Ac attracted the most number of males. The results suggested that E7Z9-12:Ac is one of the sex pheromone components and may act as a synergist.