Determination of Genetic Variation and Relationship among Zingiberaceae Species Using Polymerase Chain Reaction-Restriction Fragment Length Polymorphism Technique (PCR-RFLP)

Gan, Hui Ying (2022) Determination of Genetic Variation and Relationship among Zingiberaceae Species Using Polymerase Chain Reaction-Restriction Fragment Length Polymorphism Technique (PCR-RFLP). Final Year Project (Bachelor), Tunku Abdul Rahman University College.

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Abstract

The family of Zingiberaceae consists of more than 1200 species all over the world. Zingiber officinale which also known as ginger is widely used by people in their daily life such as spice, medicine and provide a lot of health benefits. There are various ginger cultivars tested in this study including Bentong ginger, Bara ginger, Kampung ginger, Red ginger, Red Button ginger, Tanjung Sepat ginger, Karak ginger, Thai ginger, Tambunan ginger, China ginger, Alpinia galanga and Turmeric ginger. It was found that people hard to distinguish among different cultivars by just observe the morphology. Thus, molecular technique which involve PCR-RFLP required in order to evaluate the relationship between ginger cultivars. The aim of present study is to determine the genetic variation and relationship among Zingiberaceae species using restriction fragment length polymorphism technique. Various chloroplast DNA extraction methods were investigated to choose the best chloroplast DNA extraction method through Nanodrop measurement and evaluate DNA quality by genomic and PCR gel run. It was found that liquid nitrogen sucrose density gradient centrifugation method is the best method to extract high quality chloroplast DNA. Reliable measurement of DNA concentration and purity ratio are important for this genetic study to proceed with extraction of different ginger cultivars using best extraction method. Primer set trnS-psbC and trnH-trnK were used to target certain region of chloroplast DNA of ginger cultivars using thermal cycler. The amplified products were cut with restriction enzyme HaeIII and HinfI to generate the restriction profile. The different restriction patterns allow for genetic differentiation and relationship identification among ginger cultivars. Based on the result, Red Button ginger has distinct banding patterns which differ from the other ginger species. More combination of primer sets and restriction enzymes required in order to further differentiate and classify the cultivars.