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Grass carp SERPINA1 CoIP with hepatopancreas mass spectrometry analysis

ABSTRACT: SERPINA1, a member of the serine protease inhibitor family, plays a role in viral infection and inflammation by regulating the activities of serine and cysteine proteases. To further investigate the antiviral role SERPINA1 played in GCRV (Grass carp Reovirus) infection, a polyclonal antibody of SERPINA1 was prepared, and the protein interacting with SERPINA1 was screened by CoIP/MS in grass carp hepatopancreas tissue. Samples of hepatopancreas tissues from grass carp (n=3) before (healthy) and 8 days after the infection (post-infection) were selected. Grass carp were infected by intraperitoneal injection. The total tissue proteins were extracted according to the manufacturer’s instructions for cell lysate (Beyotime, Shanghai, China). The types and abundance of proteins bound to SERPINA1 before and after the infection were detected and analyzed using CoIP-MS.

INSTRUMENT(S): TripleTOF 5600

ORGANISM(S): Ctenopharyngodon Idella

TISSUE(S): Liver

SUBMITTER: Yangyang Li

LAB HEAD: Yaping Wang

PROVIDER: PXD036356 | Pride | 2022-08-31

REPOSITORIES: Pride

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			Action	DRS
	1912052_CLM_C.wiff	Wiff
	1912052_CLM_C.wiff.scan	Wiff
	1912052_CLM_C_PeptideSummary.txt	Txt
	1912052_CLM_C_ProteinSummary.txt	Txt
	1912052_CLM_T.wiff	Wiff

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Project description:Purpose:Salinity is an important environmental factor that affects the physiological activities of fish. The goals of this study are investigating the effect of different saline-alkali stress on grass carp (Ctenopharyngodon idella). Methods: Grass carp individuals, averaging 12 cm in body length, were obtained from Duofu fish farm (Wuhan, China) and cultured at recirculating aquaculture system for 2 weeks before the experiment began. For the challenge, all grass carp were randomly divided into three groups, and then cultured at saline-alkali water with the concentration of 0, 3‰ and 6‰. After 30 days, some grass crap cultured at 3‰ and 6‰ saline-alkali water were injured. At the same time, gill samples of grass carp were collected from 0, 3‰ (grass carp was not injured), 3‰ (grass carp was injured), 6‰ (grass carp was not injured) and 6‰ (grass carp was injured)saline-alkali groups. Total RNA of all samples was isolated using TRIzol® Reagent (Invitrogen) according to the manufacturer's introduction. RNA integrity was assessed using an Agilent 2100 bioanalyzer (Agilent, USA). Samples with RNA integrity numbers (RINs) ≥ 7.5 were subjected to cDNA library construction using TruseqTM RNA sample prep Kit (Illumina). Results:A total of 15 were processed for transcriptome sequencing, generating 94.99Gb Clean Data. At least 5.76Gb clean data were generated for each sample with minimum 91.87% of clean data achieved quality score of Q30. Clean reads of each sample were mapped to specified reference genome. Mapping ratio ranged from 88.59% to 92.84%. The expression of genes was quantified and differentially expressed genes were identified based on their expression.Criteria for differentially expressed genes was set as Fold Change(FC)≥1.5 and Pvalue<0.05. Fold change(FC) refers to the ratio of gene expression in two samples. These DEGs were further processed for functional annotation and enrichment analysis. Conclusions: Our study represents Effects and molecular regulation mechanisms of saline-alkali stress on the healthy grass carp by using RNA-seqtechnology. Our results show that saline-alkali stress will impair the immune system of grass carp.

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Grass carp SERPINA1 CoIP with hepatopancreas mass spectrometry analysis

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