REAGENT or RESOURCE | SOURCE | IDENTIFIER |
---|---|---|
Chemicals, Peptides, and Recombinant Proteins | ||
BsmAI | New England Bioloabs | R0529 |
Mouth aspirator and Microcapillary tubes | Sigma | P0674 |
Levamisole hydrochloride | Sigma | L0380000 |
Trizol Reagent | Life Technologies | 15596026 |
Pronase | Sigma | 10165921001 |
30 μm filters | Sysmex | SYS-04-004-2326 |
Trizol LS Reagent | Life technologies | 10296028 |
DTT | Sigma | 10197777001 |
MiRneasy column RNA kit | QIAGEN | 217004 |
Phenol Chloroform Isoamyl | Sigma | P2069 |
Heavy Phase Lock tube | QuantaBio | 23028330 |
RNA 5′ Polyphosphatase | Epicenter | RP8092H |
NEBNext® Multiplex Small RNA Library Prep Set for Illumina | New England Biolabs | E7300 |
TruSeq Small RNA Library Prep Kit | Illumina | RS-200-0012 |
TapeStation screen tapes | Agilent | 5067-5582 5067-5588 |
TapeStation reagents | Agilent | 5067-5583 5067-5589 |
E-Gel 4% agarose | Life Technologies | G401004 |
MinElute DNA purification kit | QIAGEN | 28006 |
SMART-Seq v4 Ultra Low Input RNA Kit | Nextera | 634890 |
Nextera XT DNA Library Preparation Kit | Illumina | FC-131-1024 |
10X Phosphate Buffered Saline (PBS) | Ambion | AM9624 |
Formaldehyde | Sigma | F8775 |
RNase free Nuclease-free water | Ambion | AM9932 |
Dextran sulfate | Sigma | D8906-50G |
Escherichia coli tRNA | ROCHE | 10109541001 |
Vanadyl ribonucleoside complex | New England Biolabs | S1402S |
RNase free BSA | Ambion | AM2618 |
Formamide | Ambion | AM9342 |
20X SSC | Ambion | AM9763 |
1 M Tris-HCl, pH 8.0 | Ambion | AM9855G |
Glucose oxidase stock | Sigma | G2133 |
Catalase suspension | Sigma | C3515 |
DAPI | Sigma | D9542 |
Sodium Azide | Sigma | S2002 |
Isoamyl alcohol | Sigma | W205702 |
Benzaldehyde | Sigma | 418099 |
Diacetyl | Sigma | 8.03528 |
Butanone | Sigma | 360473 |
polydimethylsiloxane (PDMS) | Dow Corning Sylgard | 4019862 |
Milltex 1 mm biopsy punchers | Fisher | 12-460-401 |
PTFE microbore tubing | Cole-Parmer | EW-06417-21 |
Critical Commercial Assays | ||
Affymetrix GeneChip® C. elegans Genome Array oligonucleotide arrays | Thermo Fisher | 900383 |
Deposited Data | ||
Small RNA-seq data | This study | GEO: GSE124049 |
Experimental Models: Organisms/Strains | ||
wild type Bristol isolate | CGC | N2 |
rde-4(ne299) | Hobert lab | N/A |
otIs356[rab-3p(prom1)::2xNLS::TagRFP] | Oren-Suissa lab | OH10690 |
sid-1(qt9) | CGC | HC196 |
ttTi5605 II; unc-119(ed3) III; oxEx1578 | CGC | EG6699 |
rde-4(ne299) (6X outcrossed) | This study | BFF12 |
sid-1(qt9); rde-4(ne299) | This study | BFF13 |
rde-4(ne299); pigEx9[Psng-1::rde-4:SL2:yfp+Pmyo-3::Mcherry] | This study | BFF14 |
rde-4(ne299); pigEx10[Prgef-1::rde-4:SL2:yfp+Punc-122::gfp] | This study | BFF15 |
rde-4(ne299);pigSi3[cb-unc-119(+)Psng-1::rde-4:SL2:yfp] | This study | BFF16 |
rde-4(ne299); hrde-1(pig4) | This study | BFF17 |
rde-4(ne299); hrde-1(pig4); pigEx9[Psng-1::rde-4:SL2:yfp+Pmyo-3::Mcherry] | This study | BFF18 |
rde-4(ne299); sid-1(qt9); pigEx9[Psng-1::rde-4:SL2:yfp+Pmyo-3::Mcherry] | This study | BFF19 |
unc-119(ed3) III, oxTi38[cb-unc-119(+)Ppie-1::GFP] | Gift from Christian Frøkjær-Jensen | EG6089 |
hrde-1(tm1200); EG6089 | This study | BFF20 |
rde-4(ne299); EG6089 | This study | BFF21 |
rde-4(ne299); hrde-1(pig4); EG6089 | This study | BFF22 |
otIs356[rab-3p(prom1)::2xNLS::TagRFP] (3x outcrossed) | This study | BFF30 |
rde-4(ne299); otIs356[rab-3p(prom1)::2xNLS::TagRFP] | This study | BFF31 |
rde-4(ne299); pigSi3[cb-unc-119(+)Psng-1::rde-4:SL2:yfp]; otIs356[rab-3p(prom1)::2xNLS::TagRFP]. | This study | BFF32 |
sid-1(qt9); rde-4(ne299); pigSi3[cb-unc-119(+)Psng-1::rde-4:SL2:yfp]; | This study | BFF33 |
hrde-1(pig4); rde-4(ne299); pigSi3[cb-unc-119(+)Psng-1::rde-4:SL2:yfp)] | This study | BFF34 |
kyEx2373[Pstr-2::Gcamp; Punc-122:GFP] | Hendricks lab | CX10281 |
rde-4(ne299); kyEx2373 [Pstr-2::Gcamp; Punc-122:GFP] | This study | BFF35 |
rde-4(ne299); saeg-2(syb776) | SunyBiotech | PHX776 |
rde-4(ne299); saeg-2(syb777) | SunyBiotech | PHX777 |
rde-4(ne299); saeg-2(syb776) (x2 outcrossed to rde-4) | This study | BFF36 |
rde-4(ne299); saeg-2(syb777) (x2 outcrossed to rde-4) | This study | BFF37 |
rde-4(ne299); saeg-2(pig5) | This study | BFF38 |
rrf-1(ok589) | CGC | RB798 |
dcr-1(mg375) | CGC | YY470 |
ergo-1 (tm1860) | CGC | WM158 |
rrf-3(pk1426) | CGC | NL2099 |
mut-16(pk710) | CGC | NL1810 |
eri-6(mg379) | CGC | GR181 |
nrde-3 (gg66) | CGC | YY158 |
Recombinant DNA | ||
pos-1 RNAi plasmid | Vidal RNAi library | N/A |
mel-26 RNAi plasmid | Vidal RNAi library | N/A |
GFP RNAi plasmid | Hekimi lab- McGill | N/A |
Oligonucleotides | ||
PCR 1-FWD: genomic DNA rde-4 ATGGATTTAACCAAACTAACGTTTG | IDT | N/A |
PCR 1-RV: genomic DNA rde-4 TCAATCCGTGAAATCATAGGTGTTG | IDT | N/A |
PCR 2-FWD: Psng-1 AAACTGCAGTTAATTGTTAATTATCTAAGCTTG | IDT | N/A |
PCR 2-RV: Psng-1 CGGGATCCGCTAAAATAAAAGAAATATAGAGG | IDT | N/A |
PCR 3-FWD: Prgef-1 AAACTGCAGGCAGAATCGAGTCAACTGAAATCCG | IDT | N/A |
PCR 3-RV: Prgef-1 AAAGGATCCCGTCGTCGTCGTCGATGC | IDT | N/A |
PCR 4-FWD: hrde-1 (pig4) TCCACGTCCAATCCTTTGAGTG | IDT | N/A |
PCR 4-RV: hrde-1 (pig4) AAGTTGCCCAGGGGGGT | IDT | N/A |
PCR 5-FWD: saeg-2 (pig5) / saeg-2 (syb776) / saeg-2 (syb777)) TCGGCACGAAATATGTTG | IDT | N/A |
PCR 5-RV: saeg-2 (pig5) / saeg-2 (syb776) / saeg-2 (syb777) AGATAGCATCTACCGGGTGCC | IDT | N/A |
PCR 6-FWD: rde-4(ne299) CTAAGGCTGTCTATCAAAAGACGCCA | IDT | N/A |
PCR 6-RV: rde-4(ne299) AAATACCAGGTGGAAATTCAGCACTTG | IDT | N/A |
PCR -FWD: hrde-1 (tm1200) TCCACGTCCAATCCTTTGAG | N/A | |
PCR -RV: hrde-1 (tm1200) AGTGGAAAGAGTCACCACTTC | IDT | N/A |
PCR 7-FWD: pigSi3[cb-unc-119(+,Psng-1::rde-4:SL2:yfp] insert TCTCACTCGTTTAGGCTATTCC | IDT | N/A |
PCR 7-RV-1: pigSi3[cb-unc-119(+,Psng-1::rde-4:SL2:yfp] insert ACCCGATGAAATACGGATGC | IDT | N/A |
PCR 7-RV-2: pigSi3[cb-unc-119(+,Psng-1::rde-4:SL2:yfp] insert TGAGCACAATGGGAATACATCAG | IDT | N/A |
PCR 75-RV-2: pigSi3[cb-unc-119(+,Psng-1::rde-4:SL2:yfp] insertPsng-1 TGAGCACAATGGGAATACATCAG | IDT | N/A |
hrde-1 sgRNA 1: 5′ GGTGTCTATGGAAACCGAGG 3′ | IDT | N/A |
hrde-1 sgRNA 2: 5′ CATACGATTAGCCTCCTCCT 3′, | IDT | N/A |
rol-6sgRNA: 5′ GTGAGACGTCAACAATATGG 3′ | IDT | N/A |
rol-6(su1006) conversion template: 5′ TGTGGGTTGATATGGTTAAAC TTGGAGCAGGAACCGCTTCCAACCGT GTGCGCTGCCAACAATAT GGAGGATATGGAGCCACTGGTGTTCAGCCACCAGCACCAAC 3′. |
IDT | N/A |
saeg-2 sgRNA 1: CCAGCTAGCATGACTGCCACTTT saeg-2 sgRNA 2: CCACGAATGTTTAGAGTAAGTAA | SunyBiotech | N/A |
saeg-2 sgRNA 3: AGACGATGCACCATCTTCGTGT TTTAG AGCTAGA AATAGCAAGT saeg-2 sgRNA 4: TGGATCTAATGAGACGGGTAGTTT TAGAGCTAGAAATAGCAAGT saeg-2 sgRNA 5: CAATGAACGAAGT CATTCGGGTTT TAGAGCTAGAAATAGCAAGT saeg-2 sgRNA 6: GAA TGTTTAGAGTAAGTAACGTTTT AGAGCTAGAAATAGCAAGT |
IDT | N/A |
Software and Algorithms | ||
RMA algorithm with Partek Genomic suite v6.6 | Partek | N/A |
FastQC | Andrews, 2010 | N/A |
Cutadapt | Martin, 2011 | N/A |
Shortstack | Axtell, 2013 | N/A |
Butter | Axtell, 2014 | N/A |
HTSeq count | Anders et al., 2015 | N/A |
R Deseq2 | Love et al., 2014 | N/A |
FISH-quant | Mueller et al., 2013 | N/A |
Fiji | ImageJ | N/A |