>> Example 1: S. cerevisiae yeast transcription product profiling
>> Example 2: Knockout mouse transcription product profiling
>> Example 3: X ray irradiated culture cell transcription product profiling
>> Example 4: Drug stimulated culture cell transcription product profiling

>> Publications

Example 1: S. cerevisiae yeast transcription product profiling

MS-Profiling was used to analyze gene expression in four lots prepared from the same poly (A)+ RNA sample extracted from S. cerevisiae yeast. Electrophoretic images for all four lots matched, demonstrating HiCEP technology's high reproducibility (Fig. 3). Further, this method can detect 1.2X difference between gene expressions which undetectable by conventional hybridization.

Figure 3    HiCEP Technology's high reproducibility and detection sensitivity

Example 2: Knockout mouse transcription product profiling

Gene expression analysis was performed with genes from a knockout mouse. Not only were knocked out genes not expressed, but variations of expression levels of both known and unknown genes thought to be downstream were also observed (Fig. 6, Tab. 2). These results show that the expression of these approximately 500 genes is controlled either directly or indirectly by the knocked out gene.

Figure 6    Transcription product profiling in a knockout mouse

Table 2    Number of genes whose expression level varied was affected by knockout

Example 3: X ray irradiated culture cell transcription product profiling

Total RNA was extracted from X ray irradiated mouse embryo fibroblasts (MEF) and non-irradiated MEF, then gene expression analysis performed on both. Irradiation time was 6 hours. Results showed that X ray radiation resulted in increased expression of the p21 gene, which was then confirmed with RT-PCR (Fig. 7).

Figure 7    Fluctuations in p21 gene expression after X ray irradiation

Example 4: Drug stimulated culture cell transcription product profiling

A drug stimulus was administered to mouse cells after 12 hours of culturing. Total RNA was extracted after 0, 12, 24, and 32 hours of culturing and gene expression analyzed. Approximately 15,000 transcription products were detected, as were approximately 50 genes whose expression level either increased or decreased by a factor of two or more (Tab. 3). This example shows how HiCEP technology may be used effectively to search for drug stimulation marker genes.

            

Publications
1.	Taizo N, Akira F, Keiji K, Tadaaki B, Yoshihisa K, Satoshi Y. Application of HiCEP to Screening of Radiation Stress-Responsive Genes in the Soil Microarthropod Folsomia candida (Collembola). Environ. Sci. Technol. 2008, 6997-7002.
2.	Fujimori A, Yaoi T, Ogi H, Wang B, Suetomi K, Sekine E, Yu D, Kato T, Takahashi S, Okayasu R, Itoh K, Fushiki S. Ionizing radiation downregulates ASPM, a gene responsible for microcephaly in humans. BBRC. 369, 953-7 (2008).
3.	Araki R, Fukumura R, Sasaki N, Kasama Y, Suzuki N, Takahashi H, Tabata Y, Saito T, Abe M. More Than 40,000 Transcripts, Including Novel and Noncoding Transcripts, in Mouse Embryonic Stem Cells. Stem Cells. 24, 2522-8 (2006).
4.	Araki R, Nakahara M, Fukumura R, Takahashi H, Mori K, Umeda N, Sujino M, Inouye ST, Abe M. Identification of genes that express in response to light exposure and express rhythmically in a circadian manner in the mouse suprachiasmatic nucleus. Brain Res. 1098, 9-18 (2006).
5.	Fujimori A, Okayasu R, Ishihara H, Yoshida S, Eguchi-Kasai K, Nojima K, Ebisawa S, Takahashi S. Extremely low dose ionizing radiation up-regulates CXC chemokines in normal human fibroblasts. Cancer Res. 65, 0159-63 (2005).
6.	Suzuki K, Hattori A, Tanaka S, Masumura T, Abe M, Kitamura S. High-coverage profiling analysis of genes expressed during rice seed development, using an improved amplified fragment length polymorphism technique. Funct Integr Genomics. 5, 117-27 (2005).
7.	Fukumura R, Takahashi H, Saito T, Tsutsumi Y, Fujimori A, Sato S, Tatsumi K, Araki R, Abe M. A sensitive transcriptome analysis method that can detect unknown transcripts. Nucleic Acids Res. 31, e94 (2003).