title: Construction and analysis of liver suppression subtractive hybridization library of silver carp (Hypophthalmichthys molitrix) intraperitoneally injected with microcystin-LR.
abstract: Microcystin-LR (MC-LR) is the most frequently studied cyclic heptapeptide hepatotoxin produced by cyanobacteria. The toxin accumulates rapidly in the liver where it exerts most of its damage, but the molecular mechanisms behind its toxicity remain unclear. Here, suppression subtractive hybridization (SSH) was used to identify alterations in gene transcription of the silver carp (Hypophthalmichthys molitrix) after exposure to MC-LR. After hybridization and cloning, the forward and reverse subtractive cDNA libraries were obtained. At random, 150 positive clones (70 forward and 80 reverse) were selected and sequenced from the subtractive libraries, which gave a total of 88 gene fragment sequences (48 forward and 40 reverse). Sequencing analysis and homology searches showed that these ESTs represented 75 unique genes and 13 duplicates. Of the 75 unique genes, 38 shared high homology with fish genes of known functions, including immune-related genes, transporters and some involved in cell metabolism. Four sequenced genes (Fs59, Fs70, Rs2 and Rs15) were analyzed further using semi-quantitative RT-PCR. The genes from the forward library (Fs59 and Fs70) were found to be transcriptionally upregulated, while the genes from the reverse library (Rs2 and Rs15) were found to be transcriptionally downregulated. These results confirmed the successful construction of the subtractive cDNA library that was enriched for genes that were differentially transcribed in the silver carp liver challenged with MC-LR.
authors: College of Fisheries and Life Sciences, Shanghai Ocean University, Shanghai 201306, China.
authors: Xiancheng
authors: X
authors: Qu
authors: Kaiyue
authors: K
authors: Zhang
authors: Zhihui
authors: Z
authors: Cui
authors: Yong
authors: Y
authors: Zhang
authors: Jiaoyun
authors: J
authors: Jiang
authors: Long
authors: L
authors: Feng
authors: Qigen
authors: Q
authors: Liu
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D002347:Carps; D015870:Gene Expression / Q000187:drug effects; D015723:Gene Library; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D006824:Hybridization, Genetic; D008099:Liver / Q000187:drug effects*; D008387:Marine Toxins; D052998:Microcystins / Q000633:toxicity*; D012333:RNA, Messenger / Q000378:metabolism; D013489:Suppression, Genetic; D014874:Water Pollutants, Chemical / Q000633:toxicity*
chemical_list: D015415:Biomarkers; D008387:Marine Toxins; D052998:Microcystins; D012333:RNA, Messenger; D014874:Water Pollutants, Chemical; D005982:Glutathione Transferase; C057862:cyanoginosin LR