Isolation and characterisation of putative embryo gene DlRKD4 from Indonesian orchid dendrobium lineale rolfe

Abstract

In the plant model Arabidopsis thaliana, it is well known that AtRKD4 gene has a crucial function at very early stage of zygote development to begin embryo formation. The purpose of this study was to isolate and analyze the AtRKD4 homologous gene in orchids. This study found that partial gene of DlRKD4 was successfully isolated from the genome of D. lineale, possessed RWP-RK domain motif with high similarity with RKD4 protein in other orchid species. Information about the structure of AtRKD4 homologous genes in Indonesian orchid, Dendrobium lineale, will be very beneficial for orchid mass production. Method: DNA was isolated from 2-months-old protocorms (orchid embryo development), then amplified using RKD4 degenerate primers (DegRKD4). DegRKD4 primers are designed based on reference to the AtRKD4 conserved sequences, in both forward and reverse, namely DegRKD4F1 and DegRKD4R1. The PCR results showed 982 bp DNA fragment. The sequence analysis showed a 100% identity with A.thaliana RWP-RK domain-containing protein, partial mRNA, for 454 bp out of 982 bp and 94.65% identity with the 73E1-like UDP-glycosyltransferase gene from Dendrobium catenatum. The protein motif shows the conserved motif of the RWP-RK superfamily at the C-terminal of the presumed domain, showing function as nitrogen-controlled proteins that involved in plant development.