RESUMO
Quantitative real-time polymerase chain reaction is a sensitive technique for quantifying gene expression levels. One or more appropriate reference genes must be selected to accurately compare mRNA transcript levels across different samples and tissues. The freshwater pearl, Hyriopsis cumingii (Lea), is an important economic species cultured in China. To date, no reference genes for gene expression analysis in this species have been validated. This study aimed to compare the relative expression of seven housekeeping genes across different tissue types and in the mantle or pearl sac during three biomineralization processes: seasonal shell growth, shell healing and pearl-sac formation in H. cumingii. Three programs evaluated the expression stabilities of the seven genes: BestKeeper, geNorm and NormFinder. The beta actin gene (ACTB), commonly used as a housekeeping gene in many studies, was the least stable. The expressions of Ubiquitin (Ubi) and Ribosomal protein L18 (Rpl18) and Elongation factor 1-alpha (EF1α) were more stable than the remaining four genes. Therefore, we suggest that Ubi, Rpl18 and EF1α are suitable reference genes. The three selected reference genes are expected to facilitate analysis of gene expressions during shell or pearl formation in H. cumingii.
Assuntos
Exoesqueleto/crescimento & desenvolvimento , Calcificação Fisiológica/fisiologia , Regulação da Expressão Gênica , Unionidae/genética , Actinas/genética , Animais , Calcificação Fisiológica/genética , Primers do DNA/genética , DNA Complementar/química , DNA Complementar/genética , Meio Ambiente , Perfilação da Expressão Gênica , Genes Essenciais/genética , Fator 1 de Elongação de Peptídeos/genética , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Proteínas Ribossômicas/genética , Estações do Ano , Ubiquitina/genética , Unionidae/fisiologiaRESUMO
The freshwater wetland systems of India is a complex habitat that supports a broad range of diverse species including molluscs, which play an important role in supplementing third world countries. In the arsenic affected flood plains of West Bengal, a huge amount of arsenic laden groundwater is raised for the purpose of irrigation. Agricultural runoffs and flood water movement during monsoon may cause accumulation of arsenic in the adjacent freshwater aquifers, the common habitat of Lamellidens marginalis (Mollusca; Bivalvia; Eulamellibranchiata), a filter feeder, sensitive to altered environmental conditions. To examine the nature of toxicity induced by inorganic arsenic on both the haemocytes and tissues of the invertebrate heart, the animals were exposed to five different sublethal concentrations of sodium arsenite for a maximum time span of 30 days in vitro. Significant differences were recorded in the total haemocyte count, biochemical and histopathological parameters of the heart of L. marginalis under the arsenic induced stress. Our observations indicate the development of profound haematopoietic and cardiac stress under the sublethal inorganic arsenite exposure and it also implies the nature of risk imposed on the freshwater aquatic ecosystem under potential arsenic contamination.