- Title
-
Developmentally regulated gene expression of the small heat shock protein Hsp27 in zebrafish embryos
- Authors
- Mao, L., and Shelden, E.A.
- Source
- Full text @ Gene Expr. Patterns
Detection of Hsp27 and alpha-B crystallin mRNA in zebrafish embryonic and adult tissues. (A) RNA was obtained from at least 10 embryos at indicated times post fertilization and RT–PCR conducted using primers specific for Hsp27 and Ef1α mRNA (A′). The amount of starting template was adjusted to produce equal detection of ef1α. (B) Equal amounts of RNA (140 ng) obtained from embryos at 18 hpf (lane 1) and adult skeletal muscle (lane 2) analyzed for hsp27 expression. (C) Conditions and samples used to detect Hsp27 mRNA were used to examine expression of alpha-B crystallin mRNA. (D) Detection of EF1α (lane 1) and alpha-B crystallin mRNA (lane 2) in adult zebrafish lens RNA by RT–PCR. Arrows show the position of a 3Kb MW marker. EXPRESSION / LABELING:
|
Detection of hsp27 gene expression by in situ hybridization. In situ hybridization was conducted using embryos fixed 24 hpf. Embryos shown are: (A) processed without probe, (B) pretreated with DNAse I and RNAse prior to incubation with antisense, Hsp27 mRNA specific probe, (C) incubated with a sense control probe, and (D) incubated with antisense, Hsp27 mRNA specific probe. Hsp27 mRNA is detected unevenly in trunk and tail myotomes. NIH-3T3 murine fibroblasts were transiently transfected with an expression vector for an EGFP-zebrafish Hsp27 fusion protein. Cell cultures were processed using a sense control probe (E), antisense probe (F) or pretreated with DNAse and RNAse before incubation with antisense probe (G). Bar =200 μm (A–D), 100 μm (E–G). |
Detection of hsp27 expression in whole embryos during normal development. Antisense, Hsp27 mRNA specific probe (A–F,H,J,L) and a sense control probe (G,I,K) were used for hybridization. Embryos are shown at (A) high blastula, (B) 30% epiboly, (C,D):19, (E,F): 24, (G,H):36, (I,J):48, (K,L):78 hpf. Images were obtained after overnight colorimetric development, except a 1hr development was used for embryos collected at the tail bud stage (C,D). Arrows indicate the posterior tip of the embryo (C,D), midbrain/hindbrain (black arrow, H), lens (white arrow, H) and heart (J,L). Images are oriented as described in Experimental Procedures, except that the dorsal surface of the embryo is shown in E and the ventral surface is shown in C,I,J,K. Bar=300 μm. EXPRESSION / LABELING:
|
Detection of hsp27 expression in whole embryos after heat shock. Antisense, Hsp27 mRNA specific probe (B,D) and a sense control probe (A,C) were used for hybridization. Embryos were collected at 24 (A,B) and 48 (C,D) hpf and subjected to heat shock and recovery prior to fixation. Bar=300 μm. |
Detection of hsp27 expression in sectioned embryos. Hsp27 mRNA is detected in trunk myotomes of embryos at 24 hpf (A) and in embryos at 48 hpf after heat shock (C) but not in embryos without heat shock obtained at 48 hpf (B) or 54 hpf (D). Hsp27 mRNA is not detected in tissues of the head 24 hpf (E), but is detected in the lens of unstressed embryos collected at 48 (F) and 54 (H) hpf. All tissues in the heads of 48 h hpf embryos show hsp27 expression after heat stress (G). Expression was also detected in the heart of 54 hpf embryos (arrow, H) and at the midbrain/hindbrain boundary of embryos at 48 hpf (arrow, I and J). Images are transverse sections (A–H) and coronal sections (I,J). Bar =50 μm (A–D,J) and 100 μm (E–I). |
Unillustrated author statements EXPRESSION / LABELING:
|
Reprinted from Gene expression patterns : GEP, 6(2), Mao, L., and Shelden, E.A., Developmentally regulated gene expression of the small heat shock protein Hsp27 in zebrafish embryos, 127-133, Copyright (2006) with permission from Elsevier. Full text @ Gene Expr. Patterns