Ananalysis Of Transcriptome In Mangrove Clam Polymesoda Erosa Under Stress Of Polychlorinated Biphenyls (PCBs)

Polychlorinated biphenyls (PCBs) are highly toxic compounds known to cause serious organ or organ system diseases in animals, such as skin, liver, nervous system, reproductive system, and immune system, disease, etc., in addition, these chemical compounds are extremely persistent and do not break down, therefore, their introduction into the environment will be a high threat to its ecosystem. Mangrove clam Polymesoda erosa is thought to be the suitable animal for toxicological study, for this species is one of the preponderant shellfish in mangrove habitat community, does not make long distance migration and can be in a better position to absorb and concentrate pollutants. In this study mangrove clam P. erosa was used as a toxic test animal, its molecular phylogenetic status was determined through COI and 16SrRNA gene analysis, the transcriptome in the hepatopancreases and gills in the animals under the stress of PCBs were characterized by high throughput sequencing, and through softwares geNorm, NormFinder and BestKeeper to screen reference genes, real-time fluorescent quantitative PCR was carried out to confirm the preciseness of the sequencing of the transcriptome. Besides, attempt was made to look into the possibility of the applying of oxidative phosphorylation-related genes as biomarkers in pollution monitoring. Results from this study as follow:(1) The homologous sequence in COI gene from 24 Corbiculidae species, was found to cover a length of 373bp, with 150 variation locuses, accounting for 40.2% of the length; and that in 16SrRNA, cover a length of 393bp, with 110 variation locuses, accounting for 27.9%. It is obvious that gene COI contains much more genetic variation information than 16SrRNA does, and has much more potential for molecular phylogenetic analysis use. Mangrove clam P. erosa has the smallest genetic distance from Geloina erosa, only about 11.4%, and itself, together with Geloina erosa and Geloina expansa, forms a separate branch in phylogenetic tree, indicating that modern molecular phylogenetic classification was consistent with early classical works in the field, giving support to the designation of Geloina coaxans for Polymesoda erosa, or agreeing that the clam should be placed to the genus Geloina.(2) The analyses by the software geNorm showed that the expression of gene β-actin was constant most. Data from the software NormFinder analyses indicated that fi-actin gene expressed most constantly in mantle, gill and hepatopancreas tissues, but the expression in adductor muscle tissues was in a relatively low level (0.454), second only to a-tubulin (0.425). Data from BestKeeper showed that the expression of gene β-actin was most constant in mantle and gill tissues, and almost the same was true of gene GAPDH in hepatopancreas and adductor muscle tissues, but the expression of gene β-actin in these tissues was slightly low, being in second place. It was clear that β-actin could constantly express in the mantle, gill and adductor muscle tissues of the mangrove clam under the stress of PCBs, and is considered to be a suitable reference gene for qRT-PCR.(3) The results from the sequencing of transcriptome indicated that 8 target genes selected in this study enriched up-regulated expression in hepatopancreas and gill tissues, and Data from real-time fluorescent quantitative PCR were proved to be basically in agreement with the transcriptome sequencing results, and the only difference was that the multiple of up-regulation waried slightly, suggesting the sequencing of the transcriptome was reliable. It was also found that the oxidative phosphorylation- related genes in mitochondria were sensitive in response to PCBs, and might possibly become good biokarkers, but further study is needed to prove.