Insights into Early Ontogenesis of Salmo salar : RNA Extraction, Housekeeping Gene Validation and Transcriptional Expression of Important Primordial Germ Cell and Sex-Determination Genes

The challenge in extracting high-quality RNA impedes the investigation of the transcriptome of developing salmonid embryos. Furthermore, the mRNA expression pattern of important PGC and SD genes during the initial embryonic development of Salmo salar is yet to be studied. So, in the present study, w...

Full description

Bibliographic Details
Published in:Animals
Main Authors: Irfan Ahmad Bhat, Milena Malgorzata Dubiel, Eduardo Rodriguez, Zophonías Oddur Jónsson
Format: Article in Journal/Newspaper
Language:English
Published: MDPI AG 2023
Subjects:
PGCs
gonads
sex determination
housekeeping genes
S. salar
sterility
Veterinary medicine
SF600-1100
Zoology
QL1-991
Vasa
Salmo salar
Online Access:https://doi.org/10.3390/ani13061094
https://doaj.org/article/0ab6ce6f6e5e400fa1db309f4ffd7737
Description
Summary:The challenge in extracting high-quality RNA impedes the investigation of the transcriptome of developing salmonid embryos. Furthermore, the mRNA expression pattern of important PGC and SD genes during the initial embryonic development of Salmo salar is yet to be studied. So, in the present study, we aimed to isolate high-quality RNA from eggs and developing embryos to check vasa , dnd1 , nanos3a , sdf1 , gsdf , amh , cyp19a , dmrt1 and foxl2 expression by qPCR. Additionally, four HKGs ( GAPDH , UB2L3 , eEf1a and β-actin) were validated to select the best internal control for qPCR. High-quality RNA was extracted, which was confirmed by spectrophotometer, agarose gel electrophoresis and Agilent TapeStation analysis. UB2L3 was chosen as a reference gene because it exhibited lower intra- and inter-sample variation. vasa transcripts were expressed in all the developmental stages, while dnd1 was expressed only up to 40 d°C. Nanos3a was expressed in later stages and remained at its peak for a shorter period, while sdf1 showed an irregular pattern of mRNA expression. The mRNA expression levels of SD genes were observed to be upregulated during the later stages of development, prior to hatching. This study presents a straightforward methodology for isolating high-quality RNA from salmon eggs, and the resulting transcript profiles of significant PGC and SD genes in S. salar could aid in improving our comprehension of reproductive development in this commercially important species.

Similar Items