Abstract:
Horseshoe crabs are prehistoric marine arthropods belonging to the family Limulidae
and are still living today. Tachypleus gigas is one of the species of horseshoe crabs
which are still abundant in Peninsular Malaysia. The purposes of conducting this
study are to detect the DNA polymorphism among T gigas individuals and to verify
the genetic variability of this species. Genomic DNA from 33 samples were extracted
using the Wizard® Genomic DNA Purification Kit protocol. Screening of primers
were done to determine the suitable Random Amplified Polymorphic DNA (RAPD)
markers. These samples were then amplified using the Polymerase Chain Reaction
(PCR) technique. The DNA purity obtained ranged from 1.00-3.50 while the DNA
quantity ranged from 37.5-350.0 ng/μl. 40 primers were screened to test their
amplification abilities and three primers: OPC5, OPD18, and OPD20, were chosen for
further analysis. From these three primers, polymorphism was only detected using
primer OPC5 and OPD20. A total of 31 bands were obtained with 90.32%
polymorphism. The bands obtained ranged from 2-12 bands with 300-2000 bp. The
mean of the similarity index among individuals of T gigas calculated was 0.61 ± 0.16.
The genetic similarity ranged from 0.32-0.97 while the genetic distance ranged from
0. 03-1. 03. These results show that the genetic variety among individuals of Malaysian
T gigas is quite diverse as they do not share the same genetic makeup. From this
study, RAPD technique was found to be a suitable technique to generate genomic
DNA profiles of horseshoe crabs.
