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Series GSE189777 Query DataSets for GSE189777
Status Public on Dec 01, 2021
Title High-pressure sprayed siRNAs influence the efficiency but not the profile of transitive silencing
Organism Nicotiana benthamiana
Experiment type Non-coding RNA profiling by high throughput sequencing
Other
Summary In plants, small interfering RNAs (siRNAs) are a quintessential class of RNA interference (RNAi)-inducing molecules produced by the endonucleolytic cleavage of double stranded RNAs (dsRNAs). In order to ensure robust RNAi, siRNAs are amplified through a positive feedback mechanism called transitivity. Transitivity relies on RNA-DIRECTED-RNA POLYMERASE 6 (RDR6)-mediated dsRNA synthesis using siRNA-targeted RNA. The newly synthesized dsRNA is subsequently cleaved into secondary siRNAs by DICER-LIKE (DCL) endonucleases. Just like primary siRNAs, secondary siRNAs are also loaded into ARGONAUTE proteins (AGOs) to form an RNA-induced silencing complex (RISC) reinforcing the cleavage of the target RNA. Although the molecular players underlying transitivity are well established, the mode of action of transitivity remains elusive. In this study, we investigated the influence of primary target sites on transgene silencing and transitivity using the GFP-expressing Nicotiana benthamiana 16C line, high pressure spraying protocol (HPSP), and synthetic 22-nucleotide (nt) long siRNAs. We found that the 22-nt siRNA targeting the 3’ of the GFP transgene was less efficient in inducing silencing when compared to the siRNAs targeting the 5’ and middle region of the GFP. Moreover, sRNA sequencing of locally silenced leaves showed that the amount but not the profile of secondary RNAs is shaped by the occupancy of the primary siRNA triggers on the target RNA. Our findings suggest that RDR6-mediated dsRNA synthesis is not primed by primary siRNAs and that dsRNA synthesis appears to be generally initiated at the 3’ end of the target RNA.
 
Overall design siRNAs are exogenously applied on Nicotiana benthamiana via High Pressure Spraying Protocol and the transitive silencing phenotype is evaluated by sRNA-seq
 
Contributor(s) Vural Uslu V, Dalakouras A, Steffens VA, Krczal G, Wassenegger M
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Submission date Nov 29, 2021
Last update date Dec 03, 2021
Contact name Veli Vural Uslu
E-mail(s) velivural.uslu@agroscience.rlp.de
Phone 004963216711309
Organization name AgroScience GmbH AlPlanta-Institute for Plant Research
Department Epigenetics
Lab Wassenegger Lab
Street address Epigenetics, Breitenweg 71
City Neustadt-Mußbach
State/province DEUTSCHLAND
ZIP/Postal code 67435
Country Germany
 
Platforms (1)
GPL27032 Illumina NextSeq 500 (Nicotiana benthamiana)
Samples (9)
GSM5707013 siR164_1
GSM5707014 siR164_2
GSM5707015 siR164_3
Relations
BioProject PRJNA784454

Download family Format
SOFT formatted family file(s) SOFTHelp
MINiML formatted family file(s) MINiMLHelp
Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE189777_Normalized_read_counts_-_analysis_file.xlsx 41.0 Kb (ftp)(http) XLSX
GSE189777_RAW.tar 190.0 Kb (http)(custom) TAR (of XLSX)
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file
Processed data are available on Series record

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