Fig.3-4 Colony phenotypes of C100, ΔMareA and OE-MareA in different nitrogen sources media
3.2.2 Colony diameter analysis
On PDA medium, the growth of the knockout strain ΔMareA was significantly inhibited, with its colony diameter only 33.5% of that of the original strain C100. In contrast, the overexpression strain OE-MareA exhibited faster growth. This indicates that theAreA gene has an impact on the growth and development of C100. On PDA-Gln medium, all three strains showed a faster and essentially equal growth rate, suggesting that Gln as a nitrogen source can compensate for the effect of the AreA gene on C100 growth and development. However, when (NH4)2SO4was added in the PDA medium, ΔMareA strain’s ability to utilize it was significantly reduced, consistent with findings inPenicillium oxalicum (Rabha and Jha, 2018). In PDA-NaNO3 medium, ΔMareA strain exhibited poor utilization of this nitrogen source resulting in slowed down growth rate; whereas OE-MareA strain was not affected due to presence ofMareA gene and showed slightly higher growth rate than C100. In PDA-Urea medium, ΔMareA strain’s growth was higher compared to NaNO3. Overall, the MareA gene had a significant impact on monascus’ growth rate. Furthermore,the MareA gene also plays an important regulatory role in utilizing different nitrogen sources. The function of MareA gulatory varies among different fungi,which demonstrates diversity in regulatory functions performed by the MareA gene. As shown in Figure 3-5.
Fig.3-5 Colony diameters of C100, ΔMareA and OE-MareA in PDA medium with different nitrogen addition
3.2.3 Analysis of conidial production
The conidial yield of the knockout strain ΔMareA on PDA medium was significantly lower than that of the original strain C100, while the spore yield of the overexpression strain increased. The spore production of ΔMareA was relatively high on PDA-Gln medium, while it was the least on PDA-NaNO3 medium. The spore production of the overexpression strain OE-MareA was higher than that of the original strain C100 on PDA-Gln, PDA-NaNO3, and PDA-Urea media, while it was lower than that on PDA-(NH4)2SO4 media. It is speculated that a higher concentration of ammonium salt would inhibit conidial production in O E-MareA . In summary, theMareA gene affects monascus’ conidial production by inhibiting it when deleted and increasing it when overexpressed. Additionally, there is a difference in how MareA gene utilizes different nitrogen sources: Gln as a nitrogen source can offset AreA gene’s effect on conidia to some extent, while NaNO3 has the greatest impact on Monascus . As shown in Figure 3-6.