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Design and construction of single guide RNA for CRISPR/Cas9 system based on the xa13 resistance gene in some varieties of rice (Oryza sativa)

JURNAL HAMA DAN PENYAKIT TUMBUHAN TROPIKA · Vol. 23 No. 1 (2023) · hlm. 47–55

DOI
10.23960/jhptt.12347-55
ISSN
2461-0399, 1411-7525
Terbit
2023-01-18
Penerbit
Universitas Lampung
Bahasa
eng
Lisensi
creativecommons.org/licenses/by-nc/4.0

Abstrak

The xa13 gene is a recessive resistance gene against Xanthomonas oryzae pv. oryzae (Xoo) found in several rice varieties. Activation of this gene will trigger the formation of sucrose as a nutrient supply to Xoo for their growth in the plant. The disruption of this recessive gene expression in the plant can affect the negative impact of the gene, and recently can be created using clustered regularly interspaced short palindromic repeats (CRISPR) system using CRISPR-associated protein-9 (CRISPR/Cas9) technology that requires gRNA to recognize the targeted-sequence. This study aimed to design and construct the gRNA-targeting xa13 gene in rice using bioinformatics tools. CHOPCHOP was used for generated the gRNA candidates according to the target gene sequence. Two candidates of gRNA-targeted xa13 have been selected based on the analysis of bioinformatics data. Each candidate of gRNA consisted of 20 nucleotides (nt) of the target sequence upstream 3 nt of the protospacer adjacent motif (PAM) sequence (5’-NGG) targeting two exons in the xa13 gene. The gRNA1 will target exon 1 and the gRNA2 will target exon 2, with an efficiency of 52.51% and 44.63% respectively. Data showed that the GC content of all gRNA candidates ranged from 55–70% with no target-off location in the whole genome of rice. The transformation and confirmation test based on the physiological and genomic characteristics of transformants confirmed that the design has been successfully constructed.

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  • Universitas Jember
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  • Louisiana State University
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Rekan penulisOrganisasiArtikel bersama
Bambang SugihartoUniversitas Jember4
Nur Elia NadhiraAfiliasi tidak diketahui3
Sigit PrastowoUniversitas Jember2
Ankardiansyah Pandu PradanaUniversitas Jember2
Widi Kusuma WardhaniUniversitas Jember2
Zul Fauzi Nugroho HadiUniversitas Jember2
Eka DianaUniversitas Nurul Jadid1
Lusiman LusimanAfiliasi tidak diketahui1
Dinda Mala MaulidaUniversitas Jember1
Cassidy MagnonLouisiana State University1
Ahmad Ilham TanzilUniversitas Jember1
Intan Ria NelianaUniversitas Jember1
Intan Hadiatun MaghfirahUniversitas Jember1
Ummi SholikhahUniversitas Jember1
Tri Wahyu SaputraUniversitas Jember1
Weny Nailul HidayatiUniversitas Jember1
Retnosari ApriastiUniversitas Jember1
Sholeh AviviUniversitas Jember1
Galang Rizki RamadhanUniversitas Jember1
Parawita DewantiUniversitas Jember1
Syafira Fatihatul HusnaUniversitas Jember1
Erlia NarulitaUniversitas Jember1
RasmiyanaUniversitas Jember1
M Azam BaihaqiUniversitas Jember1
Desy Cahya WidianingrumUniversitas Jember1
Himmatul KhasanahUniversitas Jember1

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Sumber metadata: OAI-PMH (metadata jurnal) http://jhpttropika.fp.unila.ac.id/index.php/jhpttropika/oai, pertama di-harvest 8 Nov 2026, terakhir terlihat 8 Nov 2026.
Metadata asli (oai_dc) 
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	<dc:title xml:lang="en">Design and construction of single guide RNA for CRISPR/Cas9 system based on the xa13 resistance gene in some varieties of rice (Oryza sativa)</dc:title>
	<dc:creator xml:lang="en">Hardiyani, Wulan Arum</dc:creator>
	<dc:creator xml:lang="en">Wafa, Ali</dc:creator>
	<dc:creator xml:lang="en">Fanata, Wahyu Indra Duwi</dc:creator>
	<dc:creator xml:lang="en">Addy, Hardian Susilo</dc:creator>
	<dc:subject xml:lang="en">CRISPR/Cas9</dc:subject>
	<dc:subject xml:lang="en">gRNA</dc:subject>
	<dc:subject xml:lang="en">Oryzae sativa</dc:subject>
	<dc:subject xml:lang="en">xa13 gene</dc:subject>
	<dc:subject xml:lang="en">Xanthomonas oryzae</dc:subject>
	<dc:description xml:lang="en">The xa13 gene is a recessive resistance gene against Xanthomonas oryzae pv. oryzae (Xoo) found in several rice varieties. Activation of this gene will trigger the formation of sucrose as a nutrient supply to Xoo for their growth in the plant. The disruption of this recessive gene expression in the plant can affect the negative impact of the gene, and recently can be created using clustered regularly interspaced short palindromic repeats (CRISPR) system using CRISPR-associated protein-9 (CRISPR/Cas9) technology that requires gRNA to recognize the targeted-sequence. This study aimed to design and construct the gRNA-targeting xa13 gene in rice using bioinformatics tools. CHOPCHOP was used for generated the gRNA candidates according to the target gene sequence. Two candidates of gRNA-targeted xa13 have been selected based on the analysis of bioinformatics data. Each candidate of gRNA consisted of 20 nucleotides (nt) of the target sequence upstream 3 nt of the protospacer adjacent motif (PAM) sequence (5’-NGG) targeting two exons in the xa13 gene. The gRNA1 will target exon 1 and the gRNA2 will target exon 2, with an efficiency of 52.51% and 44.63% respectively. Data showed that the GC content of all gRNA candidates ranged from 55–70% with no target-off location in the whole genome of rice. The transformation and confirmation test based on the physiological and genomic characteristics of transformants confirmed that the design has been successfully constructed.</dc:description>
	<dc:publisher xml:lang="en">Universitas Lampung</dc:publisher>
	<dc:date>2023-01-18</dc:date>
	<dc:type>info:eu-repo/semantics/article</dc:type>
	<dc:type>info:eu-repo/semantics/publishedVersion</dc:type>
	<dc:format>application/pdf</dc:format>
	<dc:identifier>https://jhpttropika.fp.unila.ac.id/index.php/jhpttropika/article/view/660</dc:identifier>
	<dc:identifier>10.23960/jhptt.12347-55</dc:identifier>
	<dc:source xml:lang="en">Jurnal Hama dan Penyakit Tumbuhan Tropika; Vol. 23 No. 1 (2023): MARCH, JURNAL HAMA DAN PENYAKIT TUMBUHAN TROPIKA: JOURNAL OF TROPICAL PLANT PESTS AND DISEASES; 47-55</dc:source>
	<dc:source>2461-0399</dc:source>
	<dc:source>1411-7525</dc:source>
	<dc:language>eng</dc:language>
	<dc:relation>https://jhpttropika.fp.unila.ac.id/index.php/jhpttropika/article/view/660/543</dc:relation>
	<dc:rights xml:lang="en">Copyright (c) 2023 Jurnal Hama dan Penyakit Tumbuhan Tropika</dc:rights>
	<dc:rights xml:lang="en">https://creativecommons.org/licenses/by-nc/4.0</dc:rights>
</oai_dc:dc>