| Gene (Arabidopsis thaliana) |
FLAG-DCL3 |
Singh et al., 2019
|
|
Synthetic gene codon-optimized for insect cells |
| Cell line (Trichoplusia ni) |
High Five Cells in Express Five Medium |
Thermo Fisher Scientific |
Cat # B85502
|
For baculovirus replication and expression |
| Transfected construct (Escherichia coli) |
pUC57-DCL3 (synthetic DCL3, codon optimized for insect cells) |
Singh et al., 2019
|
N/A |
Strain carrying cloned FLAG-DCL3 gene |
| Transfected construct (T. ni) |
pFastBacHT B-DCL3 |
Singh et al., 2019
|
N/A |
Construct for baculovirus expression |
| Transfected construct (T. ni) |
pFastBacHT B-DCL3_E1146Q_E1329Q |
This paper |
N/A |
RNase III domain A/B double mutant |
| Transfected construct (T. ni) |
pFastBacHT B-DCL3_E1146Q |
This paper |
N/A |
RNase III domain mutant |
| Transfected construct (T. ni) |
pFastBacHT B-DCL3_E1329Q |
This paper |
N/A |
RNase III domain mutant |
| Chemical compound, drug |
PMSF |
Sigma-Aldrich |
Cat # P7626 |
Protease inhibitor |
| Chemical compound, drug |
GlycoBlue |
Thermo Fisher Scientific |
Cat # AM9515 |
|
| Chemical compound, drug |
Ribolock RNase Inhibitor |
Thermo Fisher Scientific |
Cat # EO0384 |
|
| Chemical compound, drug |
RNase Inhibitor (Murine) |
NEB |
Cat # M0314 |
|
| Chemical compound, drug |
S1 nuclease |
Promega |
Cat # M5761 |
|
| Chemical compound, drug |
Proteinase K, RNA grade |
Invitrogen |
Cat # 25530049 |
|
| Chemical compound, drug |
T4 Polynucleotide Kinase |
NEB |
Cat # M0201S |
Enzyme for end-labeling RNA |
| Chemical compound, drug |
Adenosine 5′-triphosphate magnesium salt |
Sigma-Aldrich |
Cat # A9187 |
|
| Chemical compound, drug |
[γ32P]-ATP, 6000 Ci/mmol |
PerkinElmer |
Cat #BLU002Z250UC |
Used for end-labeling RNA |
| Chemical compound, drug |
Apyrase |
NEB |
Cat # M0398L |
Hydrolyzes ATP; used in Figure 5
|
| Chemical compound, drug |
SYBR Gold Nucleic Acid Gel Stain (10,000×) |
Invitrogen |
Cat # S11494 |
|
| Chemical compound, drug |
RNA Loading Dye (2×) |
NEB |
Cat # B0363S |
|
| Chemical compound, drug |
2× TBE-Urea Sample Buffer |
Invitrogen |
Cat # LC6876 |
|
| Chemical compound, drug |
IPEGAL CA-630 |
Sigma-Aldrich |
Cat # I8896 |
Dete |
| Chemical compound, drug |
Set of rATP, rUTP, rCTP, and rGTP |
Sigma-Aldrich |
Cat # A1388 |
Used for Pol IV-RDR2 transcription |
| Chemical compound, drug |
Anti-FLAG M2 affinity gel |
Sigma-Aldrich |
Cat # A2220 |
|
| Commercial assay or kit |
Vaccinia capping system |
NEB |
Cat # M2080S |
5′ end-capping in Figure 6
|
| Software, algorithm |
Image Lab 6.0.1 |
Bio-Rad |
Cat # 12012931 |
|
| Other |
Express FiveSFM |
Thermo Fisher Scientific |
Cat # 10486025 |
Serum-free medium for High Five cells |
| Other |
Macherey-Nagel Polygram CEL 300 PEI/UV254 Polyester Sheets |
Thermo Fisher Scientific |
Cat # 10013021 |
For TLC assay of ATP hydrolysis |
