Whole transcriptome analysis on blue light-induced eye damage.

AIM: To analyze abnormal gene expressions of mice eyes exposed to blue light using RNA-seq and analyze the related signaling pathways. METHODS: Kunming mice were divided into an experimental group that was exposed to blue light and a control group that was exposed to natural light. After 14d, the mice were euthanized and their eyeballs were collected. Whole transcriptome analysis was attempted to analyze the gene expression of the eyeballs using RNA-seq to reconstruct genetic networks. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were used to reveal the related signaling pathways. RESULTS: The 737 differentially expressed genes were identified, including 430 up and 307 down regulated genes, by calculating the gene FPKM in each sample and conducting differential gene analysis. GO and KEGG pathway enrichment analysis showed that blue light damage may associated with the visual perception, sensory perception of light stimulus, phototransduction, and JAK-STAT signaling pathways. Differential lncRNA, circRNA and miRNA analysis showed that blue light exposure affected pathways for retinal cone cell development and phototransduction, among others. CONCLUSION: Exposure to blue light can cause a certain degree of abnormal gene expression and modulate signaling pathways in the eye.

A fast threshold segmentation method for froth image base on the pixel distribution characteristic.

With the increase of the camera resolution, the number of pixels contained in froth image is increased, which brings many challenges to image segmentation. Froth size and distribution are the important index in froth flotation. The segmentation of froth images is always a problem in building flotation model. In segmenting froth images, Otsu method is usually used to get a binary image for classification of froth images, this method can get a satisfactory segmentation result. However, each gray level is required to calculate each of the between-class variance, it takes a longer time in froth images with a large number of pixels. To solve this problem, an improved method is proposed in this paper. Most froth images have the pixel distribution characteristic that the gray histogram curve is a sawtooth shape. The proposed method uses polynomial to fit the curve of gray histogram and takes the characteristic of gray histogram's valley into consideration in Otsu method. Two performance comparison methods are introduced and used. Experimental comparison between Otsu method and the proposed method shows that the proposed method has a satisfactory image segmentation with a low computing time.

Construction of cDNA library of cotton mutant Xiangmian-18 library during gland forming stage.

Gossypol, a secondary metabolite stored in the glands of cotton, protecting cottonseed from consumption of human and monogastric animal. This ability is unique to the tribe Gossypieae. Although the relationship between gossypol and pigment gland has been studied for a long time, the development mechanism of pigment gland has not been investigated at molecular level. Here we described a simple and efficient method for constructing a normalized cDNA library from a cotton mutant, Xiangmian-18, during its pigments gland forming stage. It combined switching mechanism at 5'-end of RNA transcript (SMART) technique and duplex-specific nuclease (DSN) normalization methods. In a model experiment, double-stranded cDNAs were synthesized from mRNAs, processed by normalization and Sfi I restriction endonuclease, and finally the cDNAs were ligated to pDNR-LIB vector. The ligation mixture was transformed into E. coli JM109 by electroporation. Counting the number of colonies, the titer of the original library was 5.86x10(5)cfu/ml in this library. Electrophoresis gel results indicated the fragments ranged from 800bp to 2kb, with the average size of 1400bp. Random picking clones showed that the recombination rate was 94%. The results showed that the cDNA library constructed successfully was a full-length library with high quality, and could be used to screen the genes related to development of pigments gland cottons.

Molecular cloning and expression analysis of a RanBP2 zinc finger protein gene in upland cotton (Gossypium hirsutum L.).

Gossypol is an important resistant substance of Gossypium, and its storage organ is pigment gland. Although, the relationship between gossypol and pigment gland has been studied for a long time, the development mechanism of pigment gland has not been revealed up to now in molecular perspective. On the basis of differentially expressed cDNAs fragments at the stage of the cotton gland development using suppression subtractive hybridization (SSH), the complete cDNA sequence of a novel RanBP2 zinc finger protein (ZFP) gene was cloned by rapid amplification of cDNA ends (RACE) from upland cotton (Gossypium hirsutum L.), Xiangmian 18. The cotton RanBP2 ZFP cDNA (GenBank accession number: DQ173926) is 717 base pair (bp) long with an open reading frame encoding 139 amino acids, which encodes a 15.6 kDa protein. The cotton RanBP2 ZFP had three Ran-binding protein (RanBP) two zinc finger motifs and belonged to RanBP2 ZFP family. There is a 292-base non-coding sequence at 3' cDNA end, which includes polyA sequence. Sequence alignment analysis revealed that the cDNA nucleotide and its deduced amino acid sequence are moderately identical to the putative ZFP from other species. The mRNA expressing profiles of the novel ZFP gene was investigated by reverse transcription-polymerase chain reaction (RT-PCR). The result showed that it expressed at different development stages of gland, including the undeveloped stage, developing stage, developed stage and cotyledon stage. However, with the development of pigment gland, the mRNA levels in the gland-developed seed and cotyledon were increased to about 1.5 and 2 folds of that in gland-undeveloped seed, respectively, which suggested that the novel ZFP played a role in the development of the cotton gland.

[A comparative study on isoenzymes of Sarcocystis spp. from cattle and water buffaloes].

OBJECTIVE: To analyze the zymogram of peroxidase (PER) and phosphoglucose isomerase (PGI) of three species of Sarcocystis. METHODS: The collected parasites were homogenized and fragmented by ultrasonication. After centrifugation, the supernatants were analyzed by isoelectric focusing electrophoresis. RESULTS: The isolates of S. cruzi from infected water buffalo and cattle all showed identical enzyme profiles, 7 bands of PER at pH 4.44-6.98 and 6 bands of PGI at pH 4.66-6.53; and same with the isolates of S. hirsuta. 5 bands of PER at pH 4.97-7.15 and 4 bands of PGI at pH 4.70-6.51. The zymograms among S. cruzi, S. hirsuta and S. fusiformis were different considerably. CONCLUSION: The data support the hypothesis that both water buffalo and cattle are the natural intermediate hosts of S. cruzi and S. hirsuta at the gene level. S. cruzi, S. hirsuta and S. fusiformis are different species.

Characterization of the human PAP1 gene and its homologue possible involvement in mouse embryonic development.

We have identified PAP1 gene, a novel member of the immunoglobulin superfamily (IGSF) from U251-pTet-p53 cell line, which carried a wild-type p53 transgene. The gene has been localised to chromosome 16p12-13. Alignment of the predicted protein sequence for Human, Pan troglodytes, Canis, Mus musculus and Gallus gallus revealed it was highly conserved. Its homologue, IGSF6, possible involves in mouse embryonic development. The presence of IGSF6 specific transcript was detected by Northern blot in the RNAs extracted from 11 to 14 day postconception. IGSF6 expression is different in mouse embryos of the different ages. In situ hybridization performed on mice embryos sections showed the differential presence of IGSF6 in developing lung and kidney. This structure and differential expression suggests a function involvement in embryonic development, perhaps involvement in cell proliferation.

Effects of exogenous p53 transfection on the gene expression in the human brain glioma cell line U251.

The p53 gene is activated in response to several malignancy-associated stress signals by transactivation of downstream genes and by transcription-independent mechanisms. In order to identify new p53 downstream genes, we established a new system of p53 gene inducible expression, U251-pTet-p53 cell line, with the Tet-On Gene Expression System, in which exogenous p53 gene could overexpress in doxycycline (Dox) medium but not in the medium without Dox. By comparing their random primer RT-PCR products, it was proved that exogenous p53 gene expression could lead to many genes differential expression, some up-expressed and others down-expressed. All of these differential expressed genes may be p53 downstream genes. We can gain the magnitude of p53 downstream genes, which provides the basis of directly cloning of novel p53 downstream genes and further studying of p53 regulatory network.