Keragaman sekuen DNA fragmen gen penyandi ACCase subunit BCCP dari tiga tipe kelapa sawit Variability of DNA sequence of gene fragment encoding BCCP subunit of ACCase from three types of oil palm

Abstract

Summary
Heteromeric acetyl-CoA carboxylase (ht-
ACCase) is one of key enzymes in palm oil
biosynthesis. Isolation and characterization of
the gene is an important step in metabolic
engineering to increase palm oil content and
quality. The objective of this research was to
isolate DNA fragment of gene encoding biotin
carboxyl carrier protein (BCCP) subunit of ht-
ACCase from three different oil palm types
(Simalungun, Hibrida and Backcross) and
investigate the variation of its DNA sequence.
Total RNA was isolated from the mesocarp of
oil palm. DNA fragment encoding BCCP was
amplified by means of Reverse Transcriptase
Polymerase Chain Reaction (RT-PCR) using
specific primers with total RNA as a template.
The products of RT-PCR were then purified
from the gel, cloned and sequenced. The DNA
sequences were analyzed for their homologies
to BCCP gene using BlastN and aligned to
detect the sequence variability using ClustalW
program from BioEdit. The results show that
one of the two RT-PCR products at about 300
bp was highly homologous with the gene
encoding BCCP from Glycine max, Brassica
napus and Arabidopsis thaliana. Nucleotide
sequences of that BCCP fragments from the
three types of oil palm displayed some degrees
of variability. Further investigation is needed
to analyze the variability of the DNA sequences
of the full-length gene in relation with oil
content or other character

Ringkasan
Asetil-CoA karboksilase heteromerik (ht-
ACCase) merupakan salah satu enzim kunci
dalam biosintesis minyak sawit. Isolasi dan
karakterisasi gen tersebut merupakan langkah
penting dalam upaya rekayasa metabolisme
untuk peningkatan rendemen dan kualitas
minyak sawit. Penelitian ini bertujuan untuk
mengisolasi fragmen DNA penyandi subunit
biotin carboxyl carrier protein (BCCP) dari ht-
ACCase dari tiga tipe kelapa sawit yang ber-
beda (Simalungun, Hibrida dan Backcross)
dan mempelajari keragaman susunan nukleo-
tidanya. RNA total diisolasi dari mesokarp
buah sawit. Fragmen gen penyandi BCCP
diamplifikasi dengan Reverse Transcriptase-
Polymerase Chain Reaction (RT-PCR) meng-
gunakan primer spesifik dan templat RNA total.
Fragmen hasil RT-PCR dimurnikan dari gel,
diklon kemudian disekuen. Sekuen DNA yang
diperoleh dianalisis homologinya dengan gen
BCCP menggunakan BlastN dan disejajarkan
untuk mengetahui keragamannya mengguna-
kan program ClustalW dari BioEdit. Hasilnya
menunjukkan bahwa satu dari dua fragmen
hasil RT-PCR yang berukuran sekitar 300 pb
memiliki homologi yang tinggi dengan
fragmen gen penyandi BCCP dari Glycine max,
Brassica napus dan Arabidopsis thaliana.

Urutan nukleotida fragmen BCCP dari ketiga
tipe kelapa sawit menunjukkan keragaman.
Perlu analisis lebih lanjut mengenai keragaman
sekuen DNA dari gen lengkapnya dan dikaji
hubungannya dengan akumulasi minyak atau
karakter lain