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KINYA AKASHI
Nara Institute for Science and Technology

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ISOLATION, CLONING AND CHARACTERIZATION OF ACTIN-ENCODING cDNAs FROM Jatropha curcas L. IP-2P Yuniati, Ratna; Widyastuti, Utut; Sopandie, Didy; Yokota, Akiho; Akashi, Kinya; Suharsono, Suharsono
Makara Journal of Science Vol. 15, No. 2
Publisher : UI Scholars Hub

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Actin is a major component of the plant cytoskeleton, so all cells contain this protein. Actin is expressed constitutively and is involved in basic housekeeping functions required for cell maintenance. Because of this, it has been frequently used as an internal control to normalize changes in gene expressions analysis. Actually, the information of nucleotide sequence of actin gene of Jatropha curcas L. population IP-2P from Indonesia is not available yet. The objective of this research was to isolate, clone and characterize cDNA of actin genes of J. curcas IP-2P. Three partial actin gene sequences had been successfully isolated by PCR using total cDNA as template, and actin primer designed from conserved region of Arabidopsis thaliana. Nucleotide sequence analysis showed that the length of JcACT fragment is 610, 534, and 701 bp encoding 203, 177, and 234 amino acids respectively. Local alignment analysis based on mRNA sequences shows that JcACT fragment shares 98% similarity with actin mRNA of Hevea brasiliensis and 99% with actin mRNA of Ricinus communis. Based on deduced amino acid sequence, JcACT is 100% identical to actins from Prunus salicina, Gossypium hirsutum, and Betula luminifera. Even though these clones of cDNA are not completed yet, they can be used as reference in J. curcas L. gene expression analysis.
ISOLASI FRAGMEN cDNA DARI GEN PENYANDI AKTIN DARI Melastoma malabathricum Hannum, Saleha; Akashi, Kinya; Suharsono, Utut Widyastuti; Hartana, Alex
Makara Journal of Science Vol. 14, No. 2
Publisher : UI Scholars Hub

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Isolation of cDNA Fragment of Gene Encoding for Actin from Melastoma malabthricum. M. malabathricum grows well in acidic soil with high Al solubility, thereby it can be used as a model plant for tolerance to aluminum and acid stresses. Actin is housekeeping gene used as an internal control for gene expression analysis. The objective of this research was to isolate and clone the cDNA fragments of MmACT encoding for actin of M. malabathricum. Total RNA was isolated and used as the template for cDNA synthesis by reverse transcription. Four cDNA fragments of MmACT, called MmACT1, MmACT2, MmACT3, and MmACT4, had been isolated and inserted into pGEM-T Easy plasmid. Nucleotide sequence analysis showed that the size of MmACT1 and MmACT2 is 617 bp, whereas MmACT3 and MmACT4 is 735 bp. The similarity among these four MmACT is about 78%-99% based on nucleotide sequence and about 98%-100% based on amino acid sequence. Phylogenetic analysis based on amino acid sequence showed that at 1% dissimilarity, the MmACT1, MmACT2, MmACT3 and the ACT5 Populus trichocarpha are clustered in one group, while the MmACT4 is grouped with ACT9 P. trichocarpa and ACT1 Gossypium hirsutum, and these two groups are separated from actin group of monocotyledonous plants. The sequence of MmACT fragments were registered in GenBank/EMBL/DDBJ database with accession numbers AB500686, AB500687, AB500688, and AB500689.
Co-Authors AKIHO YOKOTA ALEX HARTANA Didy Sopandie Saleha Hannum Suharsono Suharsono UTUT WIDYASTUTI Utut Widyastuti Suharsono Yuniati, Ratna