Gene Cloning through PCR using Primer based of Esterase/Lipase from Brevibacillus sp.

• Main Author: Putra Rura, Chalvin
• Format: Final Project
• Language: Indonesia
• Subjects: Kimia
• Online Access: https://digilib.itb.ac.id/gdl/view/38020
• Institution: Institut Teknologi Bandung

In industrial processes, specific and stable enzymes are needed to carried various chemical reactions. Brevibacillus bacteria has the potential to produce many enzymes that are stable in many types of environment and catalyse many reactions. However there are still very few information concerning the enzymatic properties of this bacteria, that hamper its application to the food and medical industry. Exploring the chromosomal DNA of Brevibacillus thermoruber by isolating enzymes encoding gene through Polymerase Chain Reaction (shorten as PCR) may open the chance to study the bacteria from other aspects such as the protein structure and its chemical characteristic. The purpose of this research is to isolate a gene using primers derived from Brevibacillus thermoruber 423. This research is divided into 4 steps, first were designing primers, isolating chromosomal DNA from Brevibacillus thermoruber AL59, amplifying the target gene using PCR method, and sequencing of PCR product. Using primers FMA4 and RMA4 resulted in the isolation of a 668 bp gene that have a similarity to acetyl CoA carboxylase biotin (93%). The other primers, FMA4 and RMA4a, amplifies TonB-dependent receptor (75%) with a length of 1040 bp. The last pair, FCARX and RCARX – a derivative primer based on the result of FMA4-RMA4 sequence – gives a 89% similarity to anthranilate synthase component I gene. Having none of the target gene acquired indicates that the primer design is non-specific to the target gene.