cDNA Amplified from RNA of Diatom Cyclotella striata TBI Cultivated Under Various Light Conditions Nurfarida Ulfah, Sari Dewi Kurniasih Indrawan, Alfredo Kono, Yanti Rachmayanti, Zeily Nurachman*
Biochemistry Division, Faculty of Mathematics and Natural Science, Institut Teknologi Bandung, Jalan Ganesa 10, Bandung 40132, Indonesia
*zeily[at]itb.ac.id
Abstract
Lipids play a crucial role in microalgae and their metabolism was regulated by several enzymes related to their synthesis and degradation. Cyclotella striata TBI is one of tropical diatom that uses lipid as food storage component. Lipid accumulation increases when exposed to external stress factors such as high light intensity and temperature, and it could be identified from the expression at the level of RNA transcript, but RNA is an unstable compound, so cDNA synthesis is carried out as an alternative. Until now, synthesis of cDNA from diatom C. striata TBI RNA grown indoor and outdoor has not been carried out. The aim of this study was to identify cDNA by RT-PCR method from diatom C. striata TBI RNA cultivated under various light conditions. Methods of this study included cultivating C. striata TBI indoor and outdoor in the modified seawater medium, total lipid extraction, total RNA isolation, and cDNA synthesis from total RNA. The results showed that the initial C. striata TBI cell density of 4.810^{5} cells mL^{-1} increased to 2.25\times10^{6} and 1.05\times10^{6} cells mL^{-1} both indoor and outdoor cultures with cultivation for 8-9 days. This is equivalent to specific growth rates of 1.78 and 1.66 d^{-1}, or cell productivity of 2.69 and 1.52\times 10^{6} cells mL^{-1} d^{-1}, or biomass productivity of 71.27 and 47.95 mg L^{-1} d^{-1}. The total lipid content of \it C. striata TBI obtained from indoor and outdoor cultures was 32 and 49% (w/w) respectively, which equates to lipid productivity of 2.2 and 2.3 mg L^{-1} d^{-1}. RNA from \it C. striata TBI cultures both indoor and outdoor in logarithmic and stationary phases was identified by the presence of bands measuring 1400 bp, 900 bp, 250 bp which indicated 28S, 18S, and 5S RNA respectively, with concentrations ranging from 0.8 to 1.0 mg mL^{-1}. cDNA synthesis was successfully identified from the presence of internal controls bands ACTB and rbcL which were 500 bp and 750 bp, respectively. cDNA from these conditions has a concentration range of 1.0-1.6 mg mL^{-1}. cDNA from \it C. striata TBI RNA both from indoor and outdoor cultures has high purity and concentration.
Keywords: C. striata TBI, lipid, indoor, outdoor, RNA, cDNA
