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An investigation of differences in gene expression in the longissimus muscle of Meishan and Large White pigs was undertaken, using the mRNA display technique. A fragment of one differentially expressed gene was isolated and sequenced, whereupon the complete cDNA sequence was then obtained by using the rapid amplification of cDNA ends (RACE). The nucleotide sequence of the gene is not related to any known porcine gene. Sequence analysis revealed that the open reading frame of this gene encodes a protein with 322 amino acids, thus displaying high sequence identity with the PDZ binding kinase (PBK) of eleven other animal species - dog, horse, cattle, human, chimpanzee, crab-eating macaque, rhesus monkey, rat, mouse, gray short-tailed opossum and platypus, so it can be defined as the porcine PBK gene. This gene was finally assigned GeneID: 100141310. Phylogenetic tree analysis revealed that the swine PBK gene has a closer genetic relationship with the PBK gene of platypus. Gene expression analysis of eight tissues of a Meishan x Large White cross showed that the porcine PBK gene is differentially expressed in various tissues. Our experiment established the primary foundation for further research on this gene.
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Objective To explore the differentially expressed genes of gastric cancer,matched normal gastric tissue and premalignant lesions. Methods The differentially expressed cDNA bands were isolated and identified by fluorescent differential display and then reamplified by PCR.After being cloned,all cDNA fragments were sequenced. Through BLAST,the results were compared with GenBank database for homology analysis. The expression of the cDNA fragment in different tissues was confirmed by Northern blot. SMART-RACE(rapid amplication of cDNA ends) was used to amplify the full length cDNA sequence. Bioinformatics analysis was performed to analysis its function. Results A differentially expressed cDNA fragment expressed lower in gastric cancers and premalignant lesions,no homology was found in GenBank. The novel cDNA fragment was given an accession number of EST(CD454989)in GenBank. A full length cDNA sequence of 1362 bp was acquired,who coded 267 amino acids,and was homologous with BAA91562.1,whose physiology function was unknown. Conclusion A differentially expressed gene was found and it may be involved in process of the gastric cancer.
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Objective To identify the differentially expressed genes in Uigur and Kazak patients with and without type 2 diabetes mellitus.Methods The differentially expressed cDNA bands were isolated by fluorescent mRNA differential display from peripheral blood leucocyte of the Uigur and Kazak patients with type 2 diabetes mellitus and the normal controls. After being cloned, all cDNA fragments were sequenced, then underwent sequence analysis, homogenous comparison,and Northern blot analysis. Results Z5、Z8、Z15 differentially expressed cDNA fragments were found.They were over-expressed in the normal controls and were lower or scarced in the Uigur and Kazak patients with type 2 diabetes mellitus. They were selected for sequencing and hybridization. Z5、Z8 showed highly homologous to cellular repressor of E1A-stimulated genes,Z15 are unknown. Conclusion Three differentially expressed genes may have a potential relation with type 2 diabetes mellitus.
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Objective To screen and identify differentially expressed genes in colorectal carcinoma and explore the possible molecular pathogenesis of colorectal carcinoma.Methods The differentially expressed cDNA bands in colorectal carcinoma specimens and matched adjacent normal tissues were isolated by fluorescent mRNA differential display.Following differential display PCR (DD-PCR),all cDNA fragments were sequenced.By using BLAST software,the sequencing results were compared with Genebank database for homologue analysis.RT-PCR was used to detect the expression of one of the differentially expressed genes in colorectal carcinoma samples were identified by semi-RT-PCR.Results BLAST analysis revealed that the cDNA band was homologous to DDX32 (99%) and its up-regulated expression in colorectal carcinoma tissues was confirmed by RT-PCR (P
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Twenty cDNA differential fragments were isolated from the hippocampus of rats in epileptic state using mRNA differential display technique. Four fragments were sequenced and compared with the known sequences in the Genebank, which showed that ERG8, ERG11, ERG12had no significant identity to any known sequences; ERG14 had 64%-69% identity to microtubulin-associated protein of the rat. Because the differential expression of these genes was caused by epilepsy inducer coriaria lactone (CL) and anti-epilepsy drug MK-801 and ERG8 might be a novel candidate epilepsy gene; ERG11 and ERG12 might be novel candidate anti-epilepsy genes.Since the microtubulin-associated protein is closely associated with the collateral sprouting of mossy fibers in the hippocampus of seizured rat, the high expression of ERG14 in the early stage of epilepsy might predict the growth of axon and formation of synapse.
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Twenty cDNA differential fragments were isolated from the hippocampus of rats in epileptic state using mRNA differential display technique. Four fragments were sequenced and compared with the known sequences in the Genebank, which showed that ERG8, ERG11, ERG12had no significant identity to any known sequences; ERG14 had 64%-69% identity to microtubulin-associated protein of the rat. Because the differential expression of these genes was caused by epilepsy inducer coriaria lactone (CL) and anti-epilepsy drug MK-801 and ERG8 might be a novel candidate epilepsy gene; ERG11 and ERG12 might be novel candidate anti-epilepsy genes.Since the microtubulin-associated protein is closely associated with the collateral sprouting of mossy fibers in the hippocampus of seizured rat, the high expression of ERG14 in the early stage of epilepsy might predict the growth of axon and formation of synapse.
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Objective: Identification of the genes specially expressed in tumor cell but not in normal cell is important for understanding the molecular mechanisms of carcinogencsis. This study will focus on identification of differentially expressed gene fragments in human stomach cancer. Methods: By using the new developing mRNA differential display (DD) technique, genes fragments differentially expressed in stomach cancer tissues from a patient and the adjacent normal tissues beyond the tumor mass were studied. Results: Two differentially displayed complementary DNA fragments from stomach cancer tissues, scgl and scg2 (stomach cancer-associated gene, scg), cofirmed by Northern Blot, were cloned and sequenced. The nucleotide length of scgl is 194 base pairs and that of scg2 is 343 base pairs. After searching against GenBank databases by BLASTN, neither scgl nor scg2 had significant homological gene sequences with the known genes. Conclusion: These results suggested scgl and scg2 might be complementary DNA fragments of novel genes expressed in stomach cancer tissues, but not in normal tissues and may play a role in the occurrence and development of stomach cancer. Further characterization of full-length of these two complementary DNA fragments will be continued.
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Objective To screen stage\|specific expression genes of plerocercoid of Spirometra erinacei\|europaei. Methods RNA was extracted by the acid guanidinium thiocyanate\|phenol\|chloroform from plerocercoid and adult worm of Spirometra erinacei\|europaei. Contaminated DNA in the RNA was digested by RNase\|free DNase. cDNA was synthesized by using T\-\{12\}MA, T\-\{12\}MC, T\-\{12\}MG and T\-\{12\}MT primers, and PCR was then done using the same T\-\{12\}MN and one random primers. The PCR products were fractionated on 8% denatured polyacrylamide gel, differential bands of plerocercoid found in the gel were cut out, amplified by PCR and sequenced after the gel was dried out and autoradiographed. Northern hybridization was conducted to identify the stage\|specific expression genes. Results Eleven differential bands were selected from the gel and classified into 3 kinds of gene fragments by hybridization after they were amplified by PCR. The fragments 1 and 2 were confirmed to express specifically in plerocercoid by Northern hybridization, but the fragment 3 was expressed in both plerocercoid and adult worm. When the 3 gene fragments were homologically analyzed in GenBank, the sequence which was homologous with the fragments 1 and 2 was not found, but the fragment 3 had high homology with many kinds of 28S rRNA. Conclusion The gene expression of plerocercoid was different from that of adult worm probably because they live in different hosts. Two kinds of different gene fragments in plerocercoid were identified by mRNA differential display technique.
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Objective:To search for gene related to hepatocellular carcinoma ( HCC), and study its, potential significance in liver oncogenesis. Methods:The difference in mRNA expression among HCC, nontumorous liver tissue, fetal liver tissue and normal liver tissue was investigated by differential display technique. Ten hepatic tissues of HCC versus surrounding noncancerous liver tissues were examined with MRG98.2 as a probe in Dot blot analysis. Expression of vascular endothelial growth factor (VEGF)mRNA in above mentioned samples was examined with reverse transcription- polymerase chain reaction(RT~ PCR). Results:Fifty five differentially expressed cDNA fragments were isolated from HCC samples, 24 of them were present in HCC and fetal liver tissues simultaneously. Dot blot analysis showed that differential display fragment MRG98.2 expressed in 7 cases of HCC samples (7/10)but only slightly in 2 of the surrounding noncancerous hepatic tissues (2/10). The expression of VEGFmRNA was upregulation in 6/7 of HCC samples expressed MRG98. 2. Gmclusian :The activation of some genes expressed in fetal period may have an important role in the genesis of HCC. MRG98.2 may be a gene related to HCC. Its expression in HCC correlates with VEGFmRNA. MRG98.2 expression is helpful in predicting a tendency toward invasion and metastasis of HCC.
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Objective: To study the effects of chromium on glucose and lipid metabolism and gene expression in diabetic rats.Methods: Male Wistar rats were assigned to three groups:normal control(NC), alloxan-induced diabetic control group(DM), and DM with chromium supplementation group(DM+Cr). Cr 200 ?g/(kg bw?d) was supplemented orally for 60 days. At the end of the treatment, the blood glucose, lipid and serum insulin were measured, and the changes in gene expression among three groups were studied by mRNA differential display technique.Results: Blood glucose in DM+Cr group decreased significantly than that before experiment. The levels of serum TG, TC, LDL-C and AI in DM+Cr group were lower than those of DM group, while the serum HDL-C levels were higher. Serum insulin was not improved obviously in DM+Cr group. 11 cDNA fragments larger than 400 bp expressed differences in skeletal muscles between DM+Cr group and DM group and were isolated, 4 of which expressed higher in DM+Cr group, while the rest expressed higher in DM group.Conclusion: Chromium supplementation could partially improve the disorders of glucose and lipid metabolism, and have an impact on some gene expression in diabetic rats, which may contribute to the regulating effects on its disorders of metabolism.