Transfected cell microarrays essay

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Transfected cell microarrays essay

Associated features include adrenocortical proliferation, lipomas, facial angiofibromas and foregut carcinoids. The MEN1 gene maps to chromosome 11q13 and encodes a 67 kDa protein, menin 2 - 4. Mutations in the MEN1 gene frequently cause protein truncation that possibly leads to inactivated function consistent with the idea of a tumor suppressor gene 5.

Menin interacts with an array of proteins 6 - 12and is expressed in all normal tissues Loss of heterozygosity LOH at 11q13 has also been found in low frequency in sporadic tumors of the pituitary, parathyroids and endocrine pancreas, with a subset showing homozygous MEN1 mutations as well 14 - Although both in vivo and in vitro evidence of MEN1 acting as a classical suppressor is substantial 818 - 27knowledge of the molecular mechanisms and pathways associated with the MEN1 suppressor gene is limited.

The effects of MEN1 in human neuroendocrine cells and its role in gene regulation have only been investigated in a few studies thus far 2228 - 32and the actual pathways involved in tumorigenesis have not been pinpointed to this date. To further investigate possible genes of interest in tumorigenesis previous MEN1 transfection experiments on the BON1 cell line were expanded to include oligo microarray analysis.

It has been previously reported that transfection of MEN1 to a human endocrine pancreatic tumor cell line BON1 with barely detectable protein expression of menin results in growth inhibition and differential regulation of at least 4 genes.

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In the present study, RNA sampled from the same experiments as above from MEN1-transfected BON1, as well as controls, were used and subjected to RT-differential display 33 and a oligonucleotide microarray approach.

By using the differential display methodology, 3 up-regulated M1APs menin activated pathway are reported as a result of MEN1 transfection.

Corroboration of the results was achieved by using the obtained fragments as probes for Northern blotting. These genes might be of importance in MEN1 tumor development and provide insights into downstream effects of menin expression.

Transfected cell microarrays essay

Culture for mycoplasma was negative both before and after the experimental series Isolation of RNA and Northern blots. Northern blots were repeated two or three times. The PCR products from each primer combination for the three different cDNAs were denatured and loaded side by side on 0. Gels were run on an electrophoresis apparatus Life Technologies at 70 W for 2.

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Hybridization signals were quantified using a PhosphorImager Molecular Dynamics. Differentially expressed bands were cut out using sterile scalpels. Gels were re-exposed to confirm that the right band had been cut. PCR products were purified and used for direct sequencing.

None of the differentially expressed genes generated readable sequences from direct sequencing and were therefore believed to contain contaminating sequence. To obtain isolated PCR products a subcloning approach was needed.

The cloned PCR products generally contained 2 or 3 different sequences. The original primers were used for PCR to generate fragments from the different sequences.

These were used to make probes for Northern blotting to verify differential expression. The detailed protocol for the sample preparation and microarray processing is available from Pronto Systems http: Labeled cDNAs were purified to remove unincorporated dyes. Equal amounts of cy3- and cy5-labeled cDNA were mixed and the cDNA-containing hybridization solution was hybridized to the oligoarray slides.

The arrays were washed and the slides were finally dried by centrifugation at rpm for 5 min. The images were imported into GenePix Pro 5.Among these mice, half died of B-cell acute lymphoblastic leukemia, T-cell acute lymphoblastic leukemia, or a myeloproliferative neoplasm.

Moreover, miRb is likely to transform cell lines by repressing multiple mRNA, which playing a key role in myeloid differentiation and apoptosis, such as CBFB, multiple genes involved in the p53 pathway. Published in college papers, Essay Help, Research Paper Help; No Comments.

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Transfected cell microarrays essay

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the galectins differs in binding to glycans on microarrays and on cell surfaces. These differences provide the basis for unique and/or opposing biological activities.

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2. . Furthermore, screening of transfected cell microarrays resulted in the identification of several proteins, which are involved in apoptosis, cell adhesion, and the kinase-signaling pathway [38,86,87]. Transfected cell microarrays were also utilized to analyze the effects of human herpesvirus-8 on a cellular transcription factor pathway.

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