5-dye set  (Promega)

Journal: bioRxiv

Article Title: Olfactory learning primes the heat shock transcription factor HSF-1 to enhance the expression of molecular chaperone genes in C. elegans

doi: 10.1101/152736

Figure Lengend Snippet: (A) Choice index for PA14 of wild-type animals pre-exposed to the odor of either OP50 or PA14. Preference was recorded at the times indicated on the x-axis. N = 16-17 experiments of 30 animals per condition. Student’s paired t-test. *p<0.05, **p<0.01. Legends: pre-exposure conditions (choice). (B) Survival on PA14 of ‘control’ animals and animals where hsf-1 was knocked-down by RNAi. RNAi was conducted using standard methods for feeding RNAi (see Materials and Methods). N = 3 experiments of 50 animals per condition. Log-rank test. p=0. Also see Table S1. (C–E) hsp-70 (F44E5.4 / F44E5.5), hsp-16.2 (Y46H3A.3), and hsp-16.41 (Y46H3A.2) mRNA abundance measured by qRT-PCR upon exposing animals that had been trained on OP50 or PA14 odor to a lawn of PA14. Values were normalized to wild-type animals pre-exposed to OP50 odor. N = 38 (C), 12 (D), and 10 (E) experiments of 30 animals per condition. Pairwise mean comparison from linear mixed model analysis. **p<0.01, ***p<0.001. See Materials and Methods and Table S2 for complete details. (A, C, D and E) Data represent means ± S.E.M.

Article Snippet: The fixed worms were hybridized with the F44E5.4/5 Stellaris FISH Probe set labeled with Cy5 dye (Biosearch Technologies, Inc., Petaluma, CA), following the manufacturer’s instructions available online at www.biosearchtech.com/stellarisprotocols .

Techniques: Quantitative RT-PCR

Journal: bioRxiv

Article Title: Olfactory learning primes the heat shock transcription factor HSF-1 to enhance the expression of molecular chaperone genes in C. elegans

doi: 10.1101/152736

Figure Lengend Snippet: (A) Choice index of wild-type animals for PA14, following pre-exposure to the odor of either water or 2AA. Preference was recorded at the times indicated on the x-axis. N = 10 experiments of 30 animals per condition. Student’s t-test. *p<0.05, **p<0.01, ***p<0.001. (B) hsp-70 (F44E5.4 / F44E5.5) mRNA abundance measured by qRT-PCR in wild-type animals that were pre-exposed to water ‘odor’ or 2AA odor and subsequently placed on a PA14 or OP50 lawn. Values are relative to animals pre-exposed to water. N = 21 experiments of 30 animals per condition. Pairwise mean comparison from linear mixed model analysis. ***p<0.001. See Materials and Methods and Table S2 for complete details. (C) and (D) Lifespan curves of animals (C) continuously exposed to water (control) or 2AA odor, or (D) in physical contact with water-treated (control) or 2AA-treated OP50. N= 3 experiments of 50 animals per condition. Log-rank tests. No significance. See also Table S3 and Materials and Methods. (E) Survival of wild-type animals on PA14, following pre-exposure to OP50 or PA14 odor. N = 8 experiments of 50 animals per condition. Log-rank test. p<0.001. Also see Table S4; (A, B) means ± S.E.M, and (C-E) total animals across all experiments. Legends: pre-exposure conditions (choice).

Article Snippet: The fixed worms were hybridized with the F44E5.4/5 Stellaris FISH Probe set labeled with Cy5 dye (Biosearch Technologies, Inc., Petaluma, CA), following the manufacturer’s instructions available online at www.biosearchtech.com/stellarisprotocols .

Techniques: Quantitative RT-PCR

Journal: bioRxiv

Article Title: Olfactory learning primes the heat shock transcription factor HSF-1 to enhance the expression of molecular chaperone genes in C. elegans

doi: 10.1101/152736

Figure Lengend Snippet: (A) Choice index for PA14 of wild-type and tph-1 (mg280) II animals pre-exposed to the odor of PA14. Preference was recorded at times indicated (x-axis). N = 3-4 experiments of 30 animals per condition. Student’s two-sample t-test (unequal variance) *p<0.05. (B) Choice index for PA14 of tph-1 (mg280) II animals pre-exposed to the odor of PA14, compared to the choice indices in tph-1 (mg280) II animals treated with exogenous 5-HT (see Fig S3 for rescue), and pre-exposed to PA14 odor. Preference was recorded at times indicated (x-axis). N = 4 experiments of 30 animals per condition. Student’s paired t-test. *p<0.05, **p<0.01, ***p<0.001. (C) hsp-70 (F44E5.4 / F44E5.5) mRNA abundance measured by qRT-PCR upon PA14 exposure in wild-type and tph-1 (mg280) II animals pre-exposed to OP50 or PA14. Values are relative to wild-type animals pre-exposed to the odor of OP50. N = 9 experiments of 30 animals per condition. Pairwise mean comparison from linear mixed model analysis. **p<0.01 for wild-type (odor+PA14 lawn). No significance for tph-1 (mg280) II (odor+PA14 lawn). See Materials and Methods and Table S2 for complete details. (D) hsp-70 (F44E5.4 / F44E5.5) mRNA abundance measured by qRT-PCR following exposure of animals to exogenous 5-HT. Values are relative to control water-treated animals. N = 7 experiments of 20-30 animals per condition. Student’s paired t-test. *p<0.05.

Article Snippet: The fixed worms were hybridized with the F44E5.4/5 Stellaris FISH Probe set labeled with Cy5 dye (Biosearch Technologies, Inc., Petaluma, CA), following the manufacturer’s instructions available online at www.biosearchtech.com/stellarisprotocols .

Techniques: Quantitative RT-PCR

Journal: bioRxiv

Article Title: Olfactory learning primes the heat shock transcription factor HSF-1 to enhance the expression of molecular chaperone genes in C. elegans

doi: 10.1101/152736

Figure Lengend Snippet: Serotonin-mediated learning activates HSF-1 throughout the animal. (A-C) smFISH confocal micrographs showing hsp-70 (F44E5.4 / F44E5.5) mRNA and DAPI in wild-type and tph-1 (mg280) II animals pre-exposed to OP50 or PA14, and subsequently subjected to a PA14 lawn. Images are projected Z-stack images of 10 µm sections across the (A) head, (C) intestine and (E) germline of animals. Arrowheads represent hsp-70 (F44E5.4 / F44E5.5) mRNA foci. Scale bars, 10μm. (B, D and F) Quantification of number of hsp-70 (F44E5.4 / F44E5.5) foci, in projected images. N = 8-11 animals per tissue per genotype per condition, quantified from 2-3 independent experiments (see Materials and Methods). Student’s paired t-test, *p<0.05 for wild-type (OP50 odor + PA14 lawn) compared to (PA14 odor + PA14 lawn). No significance for tph-1 (mg280) II (OP50 odor + PA14 lawn) compared to (PA14 odor + PA14 lawn). (A, B, D and F) Data represent means ± S.E.M. (A) Legends: genotype; tissue.

Article Snippet: The fixed worms were hybridized with the F44E5.4/5 Stellaris FISH Probe set labeled with Cy5 dye (Biosearch Technologies, Inc., Petaluma, CA), following the manufacturer’s instructions available online at www.biosearchtech.com/stellarisprotocols .

Techniques:

Journal: bioRxiv

Article Title: Olfactory learning primes the heat shock transcription factor HSF-1 to enhance the expression of molecular chaperone genes in C. elegans

doi: 10.1101/152736

Figure Lengend Snippet: (A) HSF-1::GFP localization in germline nuclei: control animals at ambient temperature, animals on a PA14 lawn, animals pre-exposed to water-odor or animals pre-exposed to 2AA odor, and animals allowed a 30 minute recovery following pre-exposure to water-odor or 2AA odor. Arrowheads indicate HSF-1 nuclear bodies. Scale bar, 5μm. (B) Quantification of HSF-1 nuclear bodies under all conditions listed in (A). N= 33-39 nuclei per animal, 17-25 animals per condition, across 3-4 independent experiments (See materials and Methods). Student’s two-sample t-test (unequal variance). **p<0.01 for odor only. No significance for (Odor + 30 min recovery) (C) hsp-70 (F44E5.4 / F44E5.5) mRNA abundance measured by qRT-PCR following exposure of animals to the odor of water or 2AA and allowed to recover for 30 minutes on OP50 lawn before being placed on PA14 lawns. N=5-9 experiments of 30 animals per experiment. Pairwise mean comparison from linear mixed model analysis. No significance. See Materials and Methods and Table S2 for complete details. (D) Confocal micrographs of individual Z-sections showing HSF-1 localization in germline nuclei of wild-type and tph-1 (mg280) II animals pre-exposed to water-odor or pre-exposed to 2AA odor. Scale bar, 5μm (E) Quantification of HSF-1 nuclear bodies under all conditions listed in (D). N = 50-55 nuclei per animal, 8-18 animals per condition, across 2-3 independent experiments (see Materials and Methods). Student’s two-sample t-test (unequal variance) t-test. *p<0.05.

Article Snippet: The fixed worms were hybridized with the F44E5.4/5 Stellaris FISH Probe set labeled with Cy5 dye (Biosearch Technologies, Inc., Petaluma, CA), following the manufacturer’s instructions available online at www.biosearchtech.com/stellarisprotocols .

Techniques: Quantitative RT-PCR