#TITLE_ALTERNATIVE#

• Main Author: LANTI KAYAPUTRI (NIM 20505003), INDIRA
• Format: Theses
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/8220
• Institution: Institut Teknologi Bandung

Earthworm Lumbricus rubellus (Red Worm) has been used as a traditional medicine for combating stroke. Lumbrokinase produced by earthworm species of L. rubellus shows fibrinolytic activity. Lumbrokinase consists of 6 (six) cluster, which are defined as F-III-2, F-III-1, F-II, F-I-2, F-I-1, and F-I-0. The enzyme possessing serine-like-protease activity is F-III cluster. The F-III cluster are stable at 60 degrees C and has wide range of pH between 2 and 11.<p> <br /> <br /> <br /> <br /> Earthworm species of L. rubellus is very useful for human being. However it should be noticed that the amount of lumbrokinase produced from each worm is very limited, hence special technique for increasing the amount of protein, such as recombinant DNA technique, is required.<p> <br /> <br /> <br /> <br /> The aim of the research was to isolate lumbrokinase gene from local L. rubellus and to determine its nucleotide sequence. Total RNA, mRNA, and chromosomal DNA of L. rubellus have been isolated. The resulted DNA and RNA were the used as templates in PCR and RT-PCR process to produce DNA and cDNA coding lumbrokinase gene.<p> <br /> <br /> <br /> <br /> The primers used were designed based on published sequences with accession numbers of AF304199, ABO45719, and ABO45720 obtained from Gene Bank. Clustal X, DNA Star, and Gene Doc program Primer Select, Edit Seq, and Seqman, were used in the primer design.<p> <br /> <br /> <br /> <br /> Amplification process using PCR and RT-PCR methods produced DNA fragments with the size of about 300 bp. The expected size of the amplified product was between 500 and 800 bp. Electrophoregram of nucleotide sequencing analysis showed that the PCR products were a mixture of DNA fragments. These results indicate that the designed primers could not bind specifically to the targeted lumbrokinase gene. In the future it is necessary to design a degenerate primer based on amino acid residues of lumbrokinase purified from local L. rubellus.