#TITLE_ALTERNATIVE#

• Main Author: Silaban (NIM 20589048), Ramlan
• Format: Theses
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/9012
• Institution: Institut Teknologi Bandung

ABSTRACT: <br /> <br /> <br /> The cloning of penicillin acylase gene has been done from Escherichia toll B 130 chromosomal DNA in pBR 322 plasmid, and Escherichia toll HB 101 is used as a host cell. The cloning was done with several steps, including the isolation of DNA and pBR 322 plasmid, digestion by Pst 1 restriction enzyme, ligation by T-4 DNA ligase enzyme, transformation and selection. <br /> <br /> <br /> The agarose gel electrophoresis data showed that the fragments of chromosomal DNA after digestion were between 1,258 and 15,135 base pairs, and the pBR 322 plasmid gave changed from circular to linear with 4,363 base pairs. Electrophoresis of ligation product showed that there were several variations of DNA recombinant size. <br /> <br /> <br /> DNA recombinant was tranferred chemically into the host cell using calcium chloride and heat-shock, and then plated on selective media. From this, it was found that 7,500 colonies of transformant ails grew in media containing tetracycline. <br /> <br /> <br /> Selection of 2,500 colonies showed that 115 colonies grew in media containing tetracycline and dead in media containing ampicillin. Microbiological assays data using Serratia marcescens, showed that from 115 colonies tested, there were two colonies produced penicillin acylase enzyme. <br /> <br /> <br /> The activity test of penicillin acylase enzyme using Kornfeld method, showed that specific activity of crude enzyme of B 130, SILC 59 and SILC 150 are 32.5, 64.4 and 8.4 units permiligram protein, respectively.