Saccharopine dehydrogenase (EC 1.5.1.7) regulates the last step of fungal lysine biosynthesis. The gene (Fvsdh) encoding saccharopine dehydrogenase was identified and cloned from the whole genome of Flammulina velutipes. The genomic DNA of Fvsdh is 1257 bp, comprising three introns and four exons. The full-length complementary DNA of Fvsdh comprises 1107 bp with a deduced amino acid sequence of 368 residues. A 1,000-bp promoter sequence containing the TATA box, CAAT box, and several putative cis-acting elements was also identified. The results of tissue expression analysis showed that the expression level of the Fvsdh gene was higher in the pileus than in the stipe whether in the elongation or maturation stage. Further research showed that the lysine contents were 3.03 and 2.95 mg/g in maturation-pileus and elongation-pileus, respectively. In contrast, the lysine contents were 2.49 and 2.07 mg/g in elongation-stipe and maturation-stipe, respectively. To study the function of Fvsdh, we overexpressed Fvsdh in F. velutipes and found that Fvsdh gene expression was increased from 1.1-to 3-fold in randomly selected transgenic strains. The lysine contents were also increased from 1.12-to 1.3-fold in these five transformants, except for strain T3, in which the lysine contents were the same as the control. These results indicate that the expression of the Fvsdh gene can affect the lysine content of F. velutipes. K E Y W O R D SFlammulina velutipes, lysine content, saccharopine dehydrogenase

ABSTRACT. Aldolase is a key enzyme involved in glycolysis, gluconeogenesis, and the pentose phosphate pathway. To establish the expression patterns of all three aldolase isozyme genes in different tissues and during early embryogenesis in lower vertebrates, as well as to explore the functional differences between these three isozymes, the grass carp was selected as a model owing to its relatively high glucosemetabolizing capability. Based on the cDNA sequences of the aldolase A, B, and C genes, the expression patterns of these three isozymes were analyzed in different tissues and during early embryogenesis using quantitative real-time polymerase chain reaction (qRT-PCR). Sequence analysis of cDNAs indicated that aldolase A, B, and C (GenBank accession numbers: KM192250, KM192251, and KM192252) consist of 364, 364, and 363 amino acids, respectively. The qRT-PCR results showed that the expression levels of aldolase A, B, and C were highest in the muscle, liver, and brain, respectively. Aldolase A and C exhibited similar expression patterns during embryogenesis, with high levels observed in unfertilized and fertilized eggs and at the blastocyst stage, 2 J.J. Fan et al.Genetics and Molecular Research 16 (3): gmr16039234 followed by a decline and then increase after organogenesis. In contrast, aldolase B transcript was not detected during the unfertilized egg stage, and appeared only from gastrulation; the expression increased markedly during the feeding period (72 h after hatching), at which point the level was higher than those of aldolase A and C. These data suggest that the glucose content of grass carp starter feed should be adjusted according to the metabolic activity of aldolase B.