Identification of mouse mslp2 gene from EST databases by repeated searching, comparison, and assembling

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Abstract

The NPHS2 gene is expressed in podocytes and encodes the integral membrane protein called podocin, which is believed to play an important role in the renal function of glomerular filtration. Mutations in this gene can cause serious renal function disorders. In this study, we used data-mining techniques and bioinformatic tools to search for the mouse ortholog of the NPHS2-related gene. It might be valuable for future studies of renal diseases. We employed repeated cycles of searching, comparison, and assembling to extend the assembled EST sequences. The discovered gene sequence mslp2, an ortholog of the human SLP2 gene, was found to have a total length of 1253 bp with the amino acid coding region located in 32–1093 nt. It was further verified using the RT-PCR and RACE techniques to ensure its biological accuracy and then registered with the GenBank. When ClustalW was used for comparing the mslp2 and human SLP2 genes for similarities, the similarities were as high as 88% for nucleotide and 92% for amino acid sequences. In conclusion, we propose a method for rapid identification of the mouse ortholog gene from the human genome.

Introduction

Patients with serious kidney disorders need periodic dialysis or even renal transplants to get rid of abnormal renal syndromes, including protein urine, edema, etc. It was found that the NPHS2 gene is expressed in podocytes and encodes the integral membrane protein called podocin, which is a member of the stomatin protein family and is believed to play an important role in the renal function of glomerular filtration [1], [2]. Mutations in the human NPHS2 gene can cause serious renal function disorders. Expressed Sequence Tag (EST) databases are valuable resources for discovering new genes by in silico cloning [3], [4], [5], [6], [7]. In this paper, we present a method using repeated EST searching, multiple sequence comparisons, and sequence assembling to clone the mouse ortholog of the human NPHS2-related gene, which might be valuable for future studies of renal diseases.

Section snippets

Methodology

Data-mining techniques and bioinformatic tools were used to search for the mouse orthologs of human NPHS2-related genes by repeated cycles of assembling and extending EST sequences. A cycle consists of four steps: (1) searching for similar sequences from the mouse EST databases, (2) comparing and grouping the retrieved sequences with ClustalW, (3) assembling the sequences with the commercial software ContigExpress (Vector NTI Suite 6.0, InforMax Inc.), and (4) comparing the assembled contigs

Results

As shown in Table 2, 69 mouse EST sequences, which have great similarity with the human NPHS2 gene, were retrieved in the first cycle. The retrieved ESTs were clustered into nine groups after multiple sequence comparisons and were assembled into nine contigs. These contigs were further compared with NPHS2 (AJ279254) and its related genes including SLP1 (NM_004809), SLP2 (NM_013442), and Stomatin (NM_004099). Only four contigs, having a similarity greater than 70% with respect to one of these

Discussions and conclusions

In this paper, we presented a method for the identification of the mouse mslp2 gene. It is located at chromosome 4 A5 cM (NT_039284), including 1253 bp and encoding 353 amino acids, whereas the human SLP2 gene is located at 9p13.1, containing 3263 bp and encoding 356 amino acids. When ClustalW was used for comparing the mslp2 and human SLP2 genes for similarity, the similarity was as high as 88% for nucleotide (Fig. 2) and 92% for amino acid (Fig. 3) sequences.

Bioinformatics is an emerging and

Summary

Patients with serious kidney disorders need periodic dialysis or even renal transplants to prevent abnormal renal syndromes including protein urine, edema, etc. The NPHS2 gene is expressed in podocytes and encodes the integral membrane protein, namely podocin, which is a member of the stomatin protein family and is believed to play an important role in the renal function of glomerular filtration. It was found that mutations in the human NPHS2 gene can cause serious renal function disorders.

In

Acknowledgements

This work was partially funded by National Science Council of Taiwan, under Grant no. NSC93-E-2213-E-212-038.

Wen-Ling Chan received the M.S. degree in 2004 from the Department of Computer Science, National Chung Hsing University, Taichung, Taiwan. She works at the Department of Molecular Medicine, China Medical University Hospital. She is not only interested in biology but also in computer sciences. Currently, her research field is in bioinformatics, especially in the analysis of microarray data that are involved in gene expressions.

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Wen-Ling Chan received the M.S. degree in 2004 from the Department of Computer Science, National Chung Hsing University, Taichung, Taiwan. She works at the Department of Molecular Medicine, China Medical University Hospital. She is not only interested in biology but also in computer sciences. Currently, her research field is in bioinformatics, especially in the analysis of microarray data that are involved in gene expressions.

Yung-Fu Chen got his Ph.D. degree in 2002 from the Institute of Biomedical Engineering, National Cheng Kung University, Tainan, Taiwan, and MS degree in 1990 from Department of Electrical and Computer Engineering, University of Missouri-Rolla, USA. From September 2002 to August 2004 he was the Director of Computer Center and Associate Professor at Chungtai Institute of Health Sciences and Technology, Taichung, Taiwan. Since August 2004, he joined the faulty of Dayeh University as an Associate Professor at the Department of Computer Science and Information Engineering. Dr. Chen's research interests include bioinformatics and biomedical signal and image processing.

Jan-Gowth Chang earned his M.D. degree at Kaohsiung Medical University. He is the Director of the Department of Molecular Medicine, and Dean of Research and Development of China Medical University. Dr. Chang's primary research has involved the molecular diagnosis and treatment of hereditary disease. His work has been recognized by outstanding research awards by National Science Council, ROC (1992, 1998 and 2001).

Yung-Kuan Chan received his M.S. degree in computer science in 1991 from New Mexico Institute of Mining and Technology, USA. He received his Ph.D. in computer science and information engineering in 2000 from National Chung Cheng University, Chiayi, Taiwan. From 2001 to 2002, he worked as an Assistant Professor at the Department of Information Management, Chaoyang University of Technology. From August 2002 to July 2003, he was an Assistant Professor at the Department of Computer Science and Information Engineering, National Huwei Institute of Technology. Since August 2003, he is an Assistant Professor in the Department of Management Information Systems at National Chung Hsing University, Taichung, Taiwan. His research interests include image retrieval, image compression, image hiding, database systems, and information engineering.

Yen-Ping Chu is a Professor in the Department of Computer Science and the Chair of Management Information System at National Chung Hsing University, Taiwan, ROC. His research interests include high-speed networks, operating system, neural network and computer assistant learning.

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