Preliminary Study On Low-density CDNA Expression Arrays For Three Kinds Of Glycosyltransferases Gene Family In Different Tumor Cell Lines

Objective To analyze the differences of Glycosyltransferases gene expression profile in different human tumor cells with cDNA microarrays. Methods First,we established and optimized the molecular hybridization technique on the nylon membrane by dot blot assay. Based on it , we used the pp-GalNAcTs cDNA microarrays to detect the gene expression profile in tumor cell lines by chromogenic method of NBT/BCIP, we evaluated the sensitivity and feasibility of chromogenic method by comparing with the pp-GalNAcTs gene expression profile which we got before by using chemiluminescent method. By means of PCR, TOPO-TA cloning ,molecular hybridization techniques and corresponding detect system we evaluated the differential expression profiles of GlcNAc6ST,GalT and GnT glycosyltrasferase gene in different tumor cell lines. Results We established the dot blot technique on the nylon membrane,and detected the differential expression of pp-GalNAcT2 in different tumor cell lines. We found the detect system of chromogenic method of NBT/BCIP was low sensitivity in finding the differential expression of all the genes on the array, we validated the high expression level of pp-GalNAcT2 and T4 in different tumor cell lines, which may relate with the tumorigenesis and progression . We also established successfully the GlcNAc6ST, GalT and GnT glycosyltrasferase cDNA libraries.Using these libraries we created low-density microarrays for gene expression profiling,which can be used to detect different positive signals of GlcNAc6ST,GalT and GnT gene expression in human tumor cells with the chemiluminescent. The results showed the IGlcNAc6ST gene expression was high in human lung,gastric and leukemia cancer cells ,The expression of IGlcNAc6ST in neural tumor cell lines such as SHG44,PC12 could not be detected. The results also showed that,in different human tumor cells, GlcNAc6ST, GalT and GnT gene expression levels was different from each other. And in the same tumor cells,GlcNAc6ST,GalT and GnT gene differs in expression level. Conclusion GlcNAc6ST,GalT and GnT genes have different expression profiles in different tumor cells. Theremay have some relationships between the different expression profile especially thehighest expression level of IGlcNAc6ST and tumorigenesis and progression.Futher studywill be done.