Study Of The Chinese Surf Clam Mactra Chinensis Response To Cd²⁺ Based On The MRNA Transcription And Regulation Level

The Chinese surf clam(Mactra chinensis) is an economically important clam, mainly distributed in Liaoning and Shandong province. Because of coastal environmental deterioration and overfishing, the natural population of M. chinensis have considerably declined and the germplasm resources have degenerated. We use Illumina Hi-seq 2000 and Hi-seq 2500 CE to sequence the mRNA genome of gill and microRNA transcriptome of gills, and Quantity realtime-PCR(QRT-PCR) and EcoTM QRT-PCR was used to analyze the expression of functional genes and microRNA respectively, expecting to find the functional genes and microRNA response to the Cd2+ exposure. Through Illumina Hi-seq 2000, a total of 100,839 transcripts and 56,712 unigenes were obtained by data assembling from 39.9 million raw reads and 21,305 unigenes were annotated by hitting against NCBI database. According to the results of QRT-PCR, six functional genes responding to Cd2+ exposure were found. Heat shock protein 22(Hsp22) and cytochrome P450(CYP450(2C31)) were inhibited in the low concentration, and induced in the high concentration of Cd2+; thioredoxin peroxidase(TPx-A) was at normal level in low concentration, but induced in high concentration of Cd2+; glutathione peroxidase A(GPA), glutathioneperoxidase 1(GPA1) and Mn superoxide dismutase gene(MnSOD) were down-regulated when exposed to any treatment groups. Expression levels of the six functional genes response to Cd2+exposure indicated that these genes were linked to environmental stress.Through Illumina Hi-seq 2500, a total of 14,415,256 clean reads and 15,570,111 clean reads were yielded in the gill of control(S01) and experimental(S02) group respectively. A total of 14,584,077 sRNA, including 1,898,035 unique sRNA was generated, in which the S01 specific was 1,306,643 sRNA, including 1,039,738 unique sRNA and S02 was 772,379 sRNA, including 670,438 unique s RNA and the S01 & S02 was 12,505,055 sRNA, including 187,859 unique sRNA. The distribution of the sRNA length in the two library was similar, most of them were 26-27 nt. 27 nt was the most abundant length in S01, followded by 28 nt, 26 nt, and 23 nt; 26 nt was the most abundant length in S02, and followed by 27 nt, 28 nt and 23 nt. 50 miRNA was found in unique sRNA, including 38 conserved and 12 novel genes. Through the analyze of differential expression analysis, the expression of 5 miRNA was induced with significantly difference, and 17 miRNA was down regulated and 28 miRNA was up regulated without significantly diference. So the miRNA in gill of M. chinensis might involve the environmental stress. 542 target genes were yielded when the 50 miRNA were hit to transcriptome. And the target genes of differential expression mi RNA were annotated by hitting to the NCBI database, and 4 genes hit to the COG, 1 genes hit to the GO, 5 genes hit to the KEGG and 11 genes hit to the nr database. The genes hit to the NCBI database include E3 ubiquitin-protein ligase, Wnt signaling pathway and Regulator of G-protein signaling 22.