Chlorella Virus Separation And Purification Of The Chitinase Gene Cloning

An initial survey was done across the whole country. Our studies showed that Chlorella viruses are ubiquitously distributed in fresh-water bodies in China. In the eutrophic waters the liters of Chlorella virus are higher than those in the waters which are lack of organisms. The titers of this viral in waters are changing with the season which is higher in the summer and lower in the winter. Our research also showed that the Chlorella virus is very specific to its host. Through our detailed researches on 11 viral strains, we found that they share common molecular characteristics, such as an icosahedral shape, and a dsDNA genome. For the strict specification between the Chlorella virus and its host, the 11 strains all belong to Chlorlla strain NC64A viruses. But the research on the biological characters of these viruses showed that they have some differences: their size and their restriction fragmentation patterns (except BJ-1B AND BJ-2C). So, we think they are different strains which belong to the same type. Using 3% O% PEG8000 with 3% NaCI to deposit the Chlorella virus strain FJ-1, we found that the result of 7% PEG and 4% NaCI was the best one, and it do not influence the viral activity and its morphology. But the efficiency of deposition by PEG is lower than the efficiency of hypervelocity centrifuge. The reclamation rate of the former is 21% and the later is 36%. In order to PCR viral chitinase gene A181 a specific oligonucleolide primer was designed and the PBCV-1 genome DNA was used as the template. And then the PCR production was cloned on pUC19 between EcoRI site and Sail site. Through constructing its four subclone we determined the whole sequence of chifinase gene A181.