Identification and Isolation of Microsatellite

UMT 11th International Annual Symposium on Sustainability Science and Managemen t

09th – 11th July 2012, Terengganu, Malaysia

Isolation and Identification of Microsatellite Loci in Male and Female of False

Clownfish (Amphiprion ocellaris) using RAMs Primer SC6

Nor Athirah Sharif1, Rosliza Mohd Salim1, Shahreza Md. Sheriff2 and Nur Asma Ariffin3

1

Department of Biotechnology, Kulliyyah of Science, International Islamic University

Malaysia,

25200 Kuantan, Pahang, Malaysia,

2

Department of Aquaculture Science,

3

Department of Fisheries Science, Faculty of Fisheries and Aqua-Industry, University

Malaysia Terengganu, 21030 Kuala Terengganu, Terengganu, Malaysia.

Corresponding author’s e-mail: nurasma@umt.edu.my

Abstract

Microsatellite has become one of the most popular genetic markers widely used in many genetic

studies especially in fish. However, in order to apply this marker for genetic study it is crucial to

identify the microsatellite loci from a certain species. Currently information regarding microsatellite

loci in male and female of false clownfish (Amphiprion ocellaris) which are accessible for the

determination of gender is still lacking. Hence, this study was conducted to identify and isolate

microsatellite loci in male and female of false clownfish, A. ocellaris by using PCR based method. A

degenerate random amplified microsatellites (RAMs) primer SC6 was used to amplify the DNA

extracted from A. ocellaris fin tissues. The samples were then purified and cloned into pTZ57R/T

cloning vector. The positive clones were screened by using PCR amplification with the primer M13

Forward (-20). Twenty clones were sequenced and the results revealed a total number of 57

microsatellite loci were successfully isolated, where 30 loci were from male and 27 loci were from

female A. ocellaris. The length of nucleotide repeat sequences ranged between 3 to 14 repeats in

male and 3 to 15 repeats in female. (GA)14 was the longest nucleotide repeat sequence while (GT)3

was the...