Isolation and characterization of microsa tellite markers for the improvement of the local pineapple Ananas Comosus Varcomosus

Ong, Seong Siang (2008) Isolation and characterization of microsa tellite markers for the improvement of the local pineapple Ananas Comosus Varcomosus. Universiti Malaysia Sabah. (Unpublished)


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Microsatellite markers are co-dominant DNA based molecular markers. These molecular markers are extremely useful in plant genetic studies such as population analysis, genetic mapping, germplasm characterization, paternity testing, marker assisted selection, molecular breeding and species identification. This study reports the isolation and characterization of four novel microsatellite markers through the use of the 5'-anchored PCR. DNA was extracted from pineapple leaves and subsequently PCR was performed using a suitable degenerate primer, PCTI [5' -KKVRVRV (AC) 10-3'], to amplify regions of DNA containing microsatellite repeat sequences. The PCR products was then cloned into PCR2.l TOPOTA vector. The recombinant vector was then inserted into Invitrogen TOP10 competent cells through chemical transformation. Blue white screening was performed and 10 positive clones were selected for plasmid minipreparation using the alkaline lysis method. Four clones with inserts were then sequenced using the Genetic Analyzer 3130 DNA sequencer. The DNA sequences revealed the number of microsatellite loci. From there, specific primers pairs were designed for four loci. These microsatellite markers, designated as ACPC1714, ACPCT15A, ACPCT715B and ACPCTI49 were used to screen for polymorphism in a total of25 pineapples samples (five samples from Ranau, Beaufort, Tamparuli, Babagon and Kuching). Also another two microsatellite markers obtained from literature were used. Hoewever, primer for both loci were labelled with fluorescent dye, Fam and Vix. Analysis of the alleles was done by using the Gene mapper 4.0 software. The objectives of this research were achieved.

Item Type: Academic Exercise
Uncontrolled Keywords: microsatellite markers, DNA, extraction, pineapple leaves
Subjects: T Technology > TP Chemical technology
Divisions: SCHOOL > School of Science and Technology
Depositing User: Unnamed user with email
Date Deposited: 03 Aug 2016 06:16
Last Modified: 20 Nov 2017 03:41

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