根癌农杆菌介导转化红曲菌及转化子发酵性能的研究

Optimization of factors affecting Agrobacterium tumefaciens-mediated transformation of Monascus albidus and studies on fermentability of transformants

利用根癌农杆菌介导转化技术成功将潮霉素抗性基因转入发白红曲菌中，优化了抗生素浓度，发白红曲菌孢子浓度，根癌农杆菌浓度，共培养温度及时间，以及乙酰丁香酮浓度等转化条件，最终转化效率可达52个转化子／10^个红曲孢子。将转化子在含有潮霉素B的培养基继代培养5代，得到了多株稳定的转化子，对部分转化子进行PCR鉴定，结果进一步证实外源T-DNA已整合至转化子的染色体基因组中。实验中选取两株转化子对其发酵性能进行了研究，结果发现转化子发酵性能较原始菌株有较大提高，这说明T-DNA的插入影响了发白红曲菌的代谢过程。根癌农杆菌介导转化技术在发白红曲菌中的成功应用为研究红曲菌功能基因组及代谢调控提供了有力依据。

Monascus albidus was successfully transformed into Hygromycin B resistance gene by Agrobacterium tumefaciesnsmediated transformation（ATMT）.Various influencing infectors on transformation efficency were optimized including the concentration of antibiotics,the concentration of Monascus albidus spores ,cell density of Agrobacterium tumefaciens,co-cultivation time and temperature and the concentration of acetosyringone.The highest transformation frequency was about 52 transformants per 10^5 conidia.The transformants were successively incubated for five generations on the medium containing Hygromycin B.Several genetically stable transformants were obtained,some of them were selected to conduct PCR analysis,and the result further confimed that all transformants tested were arbitrarily integrated with exogenous T-DNA in genome.Two transformants were selected and studied.The results showed that thd fermentability of the two transformants was increased compared to Monascus albidus,and the insertion of T-DNA affected the metabolism of the transformants.The successful application of AIMT technology in Monascus albidus will give a powerful support to study functional genome and metablolic control of this organism.