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Naslov:CRISPR/Cas9-mediated fine-tuning of miRNA expression in tetraploid potato
Avtorji:ID Lukan, Tjaša (Avtor)
ID Veillet, Florian (Avtor)
ID Križnik, Maja (Avtor)
ID Coll Rius, Anna (Avtor)
ID Mahkovec Povalej, Tjaša (Avtor)
ID Pogačar, Karmen (Avtor)
ID Stare, Katja (Avtor)
ID Chauvin, Laura (Avtor)
ID Chauvin, Jean-Eric (Avtor)
ID Gruden, Kristina (Avtor)
Datoteke:URL URL - Izvorni URL, za dostop obiščite https://academic.oup.com/hr/article/doi/10.1093/hr/uhac147/6623516
 
.pdf PDF - Predstavitvena datoteka, prenos (2,94 MB)
MD5: 2AF8FACAE0FBECAECB7F6677F19FEAD0
 
Jezik:Angleški jezik
Tipologija:1.01 - Izvirni znanstveni članek
Organizacija:Logo NIB - Nacionalni inštitut za biologijo
Povzetek:MicroRNAs (miRNAs) are small noncoding RNAs, which modulate the abundance and spatiotemporal accumulation of target mRNAs at transcriptional and post-transcriptional levels and through that play important roles in several biological processes in plants. Here we show that in polyploid species, CRISPR/Cas9 system can be used for fine-tuning of miRNA expression, which can have broader range of applications compared to knock-out mutants. We established the complete pipeline for CRISPR-Cas9-mediated modulation of miRNA expression in potato. It consists of (1) design and assembly of dual sgRNA CRISPR/Cas9 constructs, (2) transient transfection of protoplasts following fast and efficient screening by high resolution melting analysis to select functional sgRNAs, and (3) stable transformation of potato explants with functional sgRNAs and selection of regenerated transgenic lines with desired mutations and desired miRNA abundance based on sequencing and RT-qPCR. We show that miRNA-editing using dual sgRNA approach results in different types of mutations among transgenic lines but also in different alleles of the same plant, which are target site-dependent. The most frequent were short deletions, but we also detected 1-nt insertions (T or G), deletions between two sgRNAs and larger deletions. miRNA abundance correlates with the frequency and type of introduced mutations, as more extensive mutations in more alleles result in lower miRNA abundance. Interestingly, some mutated loci can generate alternative miRNAs, now novel targets were however predicted for those. In all transgenic lines with Cas9 expression, we detected mutations, suggesting high efficiency of Cas9-editing. We confirmed the miRNA-editing efficiency of our optimised approach in two different potato genotypes and three different loci.
Ključne besede:CRISPR/Cas9, hypersensitive response (HR)-conferred resistance, immune signaling, live cell imaging, Solanum tuberosum (potato), spatiotemporal analysis, stromules, virus resistance
Status publikacije:Objavljeno
Verzija publikacije:Objavljena publikacija
Datum objave:30.06.2022
Leto izida:2022
Št. strani:str. 1-15
Številčenje:Vol. 9
PID:20.500.12556/DiRROS-19442 Novo okno
UDK:57
ISSN pri članku:2052-7276
DOI:10.1093/hr/uhac147 Novo okno
COBISS.SI-ID:121833731 Novo okno
Opomba:Nasl. z nasl. zaslona; Opis vira z dne 19. 9. 2022;
Datum objave v DiRROS:17.07.2024
Število ogledov:526
Število prenosov:298
Metapodatki:XML DC-XML DC-RDF
:
LUKAN, Tjaša, VEILLET, Florian, KRIŽNIK, Maja, COLL RIUS, Anna, MAHKOVEC POVALEJ, Tjaša, POGAČAR, Karmen, STARE, Katja, CHAUVIN, Laura, CHAUVIN, Jean-Eric in GRUDEN, Kristina, 2022, CRISPR/Cas9-mediated fine-tuning of miRNA expression in tetraploid potato. Horticulture research [na spletu]. 2022. Vol. 9, p. 1–15. [Dostopano 4 april 2025]. DOI 10.1093/hr/uhac147. Pridobljeno s: https://dirros.openscience.si/IzpisGradiva.php?lang=slv&id=19442
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Gradivo je del revije

Naslov:Horticulture research
Skrajšan naslov:Hortic. res.
Založnik:Nature Publishing Group in partnership with Nanjing Agricultural University
ISSN:2052-7276
COBISS.SI-ID:523115289 Novo okno

Gradivo je financirano iz projekta

Financer:ARIS - Javna agencija za znanstvenoraziskovalno in inovacijsko dejavnost Republike Slovenije
Številka projekta:P4-0165-2022
Naslov:Biotehnologija in sistemska biologija rastlin
Financer:ARIS - Javna agencija za znanstvenoraziskovalno in inovacijsko dejavnost Republike Slovenije
Številka projekta:J4-2544-2020
Naslov:Ciljana mutageneza s CRISPR/CAS9 za odpornost vinske trte in krompirja proti fitoplazmam
Financer:ANR - French National Research Agency
Program financ.:French National Research Agency (ANR)
Številka projekta:11-BTBR-0001
Naslov:Ingénierie cellulaire : amélioration et innovation technologiques pour les plantes d'une agriculture
Akronim:GENIUS
Financer:Drugi - Drug financer ali več financerjev
Program financ.:European Cooperation in Science and Technology
Številka projekta:CA18111
Naslov:COST Action PlantEd
Financer:Drugi - Drug financer ali več financerjev
Program financ.:Institut Carnot Plant2Pro program
Akronim:POTATOCRISP

Licence

Licenca:CC BY 4.0, Creative Commons Priznanje avtorstva 4.0 Mednarodna
Povezava:http://creativecommons.org/licenses/by/4.0/deed.sl
Opis:To je standardna licenca Creative Commons, ki daje uporabnikom največ možnosti za nadaljnjo uporabo dela, pri čemer morajo navesti avtorja.

Sekundarni jezik

Jezik:Slovenski jezik
Ključne besede:CRISPR/Cas9, small RNA rearrangement, potato, polyploids


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