No abstract

Cloning and protein expression in heterologous systems are very useful tools for the study of viral proteins. In this work, an in vivo cloning strategy was applied using the yeast Saccharomyces cerevisiae, as an efficient and low-cost method to clone several cDNAs from the tilapia lake virus (TiLV). Samples of infected tilapia Oreochromis niloticus tissues were taken and used to isolate their RNA and to obtain and clone the ten viral cDNAs in a shuttle plasmid. The cloning efficiencies range from 5 to 100% but for seven of the cDNAs the values were above 40%, demonstrating the high efficiency of the method.plasmid construction, Saccharomyces cerevisiae in vivo cloning, viral recombinant proteins | INTRODUCTIONIn aquaculture, the most common causes of infectious diseases are bacteria (55%) followed by viruses (23%), parasites (19%), and fungi (3%) (Asencios et al., 2016;Kibenge & Godoy, 2016). Due to the high susceptibility of the aquatic animals and the limited availability of therapeutics, viruses are the main pathogens worldwide. In the case of tilapia O. niloticus, only a few viruses have been reported to cause severe damage (Surachetpong et al., 2017).Accordingly, it is of great interest to study the tilapia lake virus (TiLV), a virus associated with massive mortality in almost 40 countries (Dong et al., 2017;Jansen & Mohan, 2017) since 2009, and was formally described for the first time in 2014 (Eyngor et al., 2014).

The cell cycle is defined as the periodic occurrence of events that are coordinately driven by the cyclin‐dependent kinases (CDKs). Cyclins are structurally related proteins whose levels fluctuate throughout the cell cycle. These CDK activators also impart distinct substrate specificity to CDKs for the temporal regulation of cell division.

The cell cycle is defined as the periodic occurrence of events that result in chromosome duplication ( deoxyribonucleic acid, DNA replication in S phase) and separation (mitosis). This process is directly regulated by both external stimuli (such as nutrient availability) and internal stimuli (such as cell size and DNA integrity). These events are co‐ordinately driven by the cyclin‐dependent kinases (CDKs) . Although the expression of CDKs typically remains relatively constant, their activities are highly regulated by CDK‐binding proteins known as Cyclins . Cyclins are structurally related proteins whose levels fluctuate throughout the cell cycle. Cyclin levels in the cell are dynamically regulated through tight control over both their rate of synthesis and degradation via ubiquitin ‐mediated proteolysis. These CDK activators also impart distinct substrate specificity to CDKs for the temporal regulation of cell division. Key Concepts: Cyclin levels oscillate throughout the cell cycle. Cyclins bind to CDKs and regulate their kinase activity. Cyclin‐CDK complexes direct the specificity of appropriate substrate interactions during the cell cycle. Inappropriate regulation of cyclins can be both the cause and the result of oncogenesis. As well as controlling normal cell cycle progression, cyclins function also in the exiting of the cell cycle (such as G0 in terminally differentiated cells and the onset of senescence caused by DNA damage).