Using modified whole-mount in situ hybridization to study mpo expression in zebrafish
- Authors:
- Li-Jing Shen
- Lan-Fang Cao
- Fang-Yuan Chen
- Yong Zhang
- Ji-Hua Zhong
- Hua Zhong
View Affiliations
Affiliations: Department of Pediatrics, Renji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200001, P.R. China, Department of Hematology, Renji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200001, P.R. China
- Published online on: February 5, 2013 https://doi.org/10.3892/etm.2013.947
-
Pages:
1043-1047
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Abstract
In this study, we cloned the myeloperoxidase (mpo) gene of zebrafish and prepared a digoxigenin-labeled mpo RNA probe to investigate mpo gene expression in zebrafish during embryonic development by whole-mount in situ hybridization (WISH). The earliest mpo expression was detected in cells of the intermediate cell mass (ICM) at 18 h post-fertilization (hpf). It was detected 1 to 2 h later in cells in the rostral blood island (RBI) and strong signals were observed in the anterior ICM. Then, it spread over the yolk sac. By 72 hpf these mpo-expressing cells were in the circulation and distributed throughout the embryo. We identified that the level of mpo expression detected by WISH at an early stage was consistent with the data of cytological analyses of adult fish. The use of this method enabled us to track the gene changes that took place before morphological phenotypes were detected, as well to as investigate the hematopoietic cell fate in mutational or transgenic models in vivo. In this study, we modified several steps of WISH. The improved hybridization results demonstrated high specificity, distinct coloration and low background figures.
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