Isolation and characterization of near complete cDNA and genomic sequences coding for granule-bound starch synthase in sago palm (Metroxylon sagu)

Lau, Jennifer Siew Kee (2001) Isolation and characterization of near complete cDNA and genomic sequences coding for granule-bound starch synthase in sago palm (Metroxylon sagu). Masters thesis, Universiti Malaysia Sarawak (UNIMAS).

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Abstract

This study involved the isolation of a eDNA and a near oomplete genomic sequence that determine the granule-bound starch synthase {GBSS) gene from sago palm (Metroxylon sagu). The eDNA was isolated via a reverse transcription-polymerase chain reaction (RT -PCR) technique using primers designed based on the conserved regions of the GBSS. The RT-PCR product obtained, 917 bp in size, was cloned and sequenced. Homology search by using the BLASTX search program confirmed that the RT-PCR product consists of part of the GBSS gene, with the deduced amino acid sequence sharing 68-7:Z01o identities with GBSS of rice, potato, maize, wheat and cassava. A pair of specific internal primers was then designed based on the RT-PCR product. These primers were used to amplify the corresponding genomic DNA region via a genomic PCR method. A 2,008 bp genomic PCR product was generated by this primer set. Cloning and sequencing of the genomic PCR product confinned that it consists of a partial genomic GBSS gene. This genomic PCR product was then used as a probe in an attempt to isolate the entire genomic locus ofthe sago GBSS via Southern hybridization.

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