Indonesian

Abstract

Semiarti E, Rozikin. 2015. Characterization of High Temperature Resistance gene (HSP70) of Vanda tricolor var. suavis forma Merapi orchids. Pros Sem Nas Masy Biodiv Indon 1: 404-408. Vanda tricolor Lindl.var.suavis Forma Merapi is an Indonesian local orchid that grows on the slopes of Mount Merapi and became the mascot of Province of Special Territory of Yogyakarta. The ability to survive on the slopes of Merapi, which is often hit by pyroclastic flows when volcano eruption occurs show the resistance to high temperatures, which is probably caused by the role of Heat Shock Protein (HSP) as molecular chaperones. The aim of this study was to isolate and characterize the HSP70 gene in V. tricolor Forma Merapi. The methods of research conducted by isolating HSP70 cDNA from cDNAlibrary of V.tricolor leaves, 2 years of age, using two sets of degenerate primers HSP70F1R1 and HSP70F2R2, that generating approximately 600 bp HSP70cDNA. The amplified cDNAs were sequenced and analyzed by the Basic Local Alignment Search Tool (BLAST), Bioedit and analyzed for a phylogenetic tree with MEGA5 and analyzed for protein motifs using online software MOTIF Search ( http://www.genome.jp/tools/motif/). The results showed that HSP70cDNA were amplified using HSP70F1R1 and HSP70F2R2 with lengths of 600 bp and 680 bp, respectively. The sequence alignment of PCR products generates cDNA with a length of 1212 bp. Phylogenetic analysis based on the amino acid sequence similarity with HSP70cDNA of other organisms showed 82-85% sequence identities, including Zea mays (82%), Dendrobium officinale (84%), Arabidopsis lyrata (84%) and Malus domesticus (85%). In HSP70 V. tricolor there are conserved domains for all types of HSP, i.e sugar - nucleotide binding site actin HSP70 kinase superfamily in the a.a sequence 1-400, Pox Ag35 superfamily at a.a sequences 750-110 and NAD - GH a.a 250- 800. The third domain is common for HSP70 proteins with several variations. V.tricolor HSP70 having a sequence with the sequence-specific pattern of HSP70 is [ LIVMY ] -x- [ LIVMF ] -xGGx- [ ST ] - { LS } - [ LIVM ] -Px- [ LIVM ] -x- [ DEQKRSTA ] at a.a sequences 96-110. Phylogenetic analysis by comparing protein HSP70 V. tricolor with protein HSP70 from other organisms showed that V. tricolor has a specific amino acid that is a characteristic protein of HSP70 V.tricolor, which might support the plant to survive in a habitat with high temperatures.