The Identification Of Chlorella Sp., Selection Of Prouctive Strains And Optimization Of Culture Conditions

In this paper, the alge was preliminary identificated as Chlorella sp. based on its morphological features. Then the Chlorella sp. was identificated as Chlorella pyrenoidosa using molecular methods. The monoclony isolation and ultraviolet radiation mutagenesis are used to select productive strains which grow faster and contain more protein. The culture conditions such as light, tempreture and nitrogen source of productive mutants was optimized.The ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit (rbcL) gene was cloned to identificated Chlorella sp.. we analysised the sequence using BLASTn and molecular systematics methods. The result showed that the Chlorella sp. and Chlorella pyrenoidosa have the closest relationship with high bootstrp values(100%). It can be concluded that the Chlorella sp. is Chlorella pyrenoidosa.We filtrated a strain, named Chlorella pyrenoidosa Q7, by single clone method. Then the Chlorella pyrenoidosa Q7 was radiated in 30W ultraviolet light. The irradiation distance was 15 cm and time was 20 min. We got 3 strains of mutants, named Chlorella pyrenoidosa M51, Chlorella pyrenoidosa M59 and Chlorella pyrenoidosa M73, from 120 clones. They grow faster and contain more protein than other strains. Compared with parent strain Q7, the growth rate of mutants M51, M59 and M73 was increased by 6.23%, 3.8% and 5.92% respectively and their protein content was increased by 2.5%, 3.1% and 1.9% respectively. Their genetic stability was analysed also. Good genetic stability was concluded.The most suitable foster situation of productive mutant strains was studied. It showed that the productive strains have biggest growth rate and bio-mass at 140μmoLm-2s-1, 25℃. The growth rate of M51 is faster than M59 and M73. So we think M51 has the potency of largescale cultivation. Then the effect of different concentration and different nitrogen sources (urea, NH4HCO3, NaNO3) on the growth and protein content of Chlorella pyrenoidosa M51 was investigated. When nitrogen sources is urea or NaNO3, the algae can not grow fast or get high bio-mass. When the NH4HCO3 is nitrogen sources, the algae can grow rapidly and its cell density can arrived 32.15×106 cell/ml. The orthogonal design is based on the results of single nitrogen source experiment. The orthogonal experiment result revealed that the optimum nitrogen group for growth is urea 700 mg/L, NH4HCO3 125 mg/L, NaNO3 0 mg/L and the optimum nitrogen group for protein content is urea700 mg/L, NH4HCO3 500 mg/L, NaNO3150 mg/L.