Lipase Synthesis Lab Report

The lipase gene sequence from Alcaligenes sp. JG3 was translated into amino acid sequence using two methods, manual translation and ExPASy online software. By manual method, amino acid sequence was obtained via codon translation one by one as shown in Figure IV.7. The sequence reference used is the lipase from A. faecalis MOR02 due to having high similarity with the lipase sequence from strain JG3 during CLUSTAL alignment and the alignment of the deduced amino acid with the referred sequence was carried out consecutively (Figure IV.8).

On the other hands, the possible ORFs were automatically obtained (Appendix 4.A) via ExPASy online software, the second method. To find the most possible one, the obtained ORFs were aligned with lipase encoding ORF from Alcaligenes faecalis resulted one sequence containing 199 aa (amino acid) that has high identity (98%) with the reference sequence as shown in Figure. IV.9. Notwithstanding, the 199 aa is shorter than A. faecalis has. As mentioned above that DF1 and DR1 amplified 1067 bp, it should encode more than 350 aa of ORF. Originally, the 1067 bp sequence encoded the same length. However, several stop codons were found along it due to poor sequencing results with lower quality value (QV) in electrophoregram. It is caused by either overlapping or broadening peaks that the representative base could not be read properly.
Conversely, since the ExPASy program using certain algorithms to convert DNA sequence into amino acid where the deletion of several bases is not allowed, other sequence references were needed to complete the amino acid encoding lipase