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      Expression Pattern in a Modified Equalized Kidney cDNA Library of Hypertensive Rat

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          Background: Genes with important functions and rarely expressed would probably more easily be cloned from a modified equalized kidney cDNA library for further investigation. Methods: A kidney cDNA library of a spontaneously hypertensive rat was synthesized by a modified equalization method. Inserts of random clones were amplified by PCR and sequenced. Sequences were compared against a nonredundant database in GenBank. The cDNA profile was compared with an expression profile of a mouse renal proximal tubule cDNA library. Seven clones were analyzed by Northern blot analysis. The cDNA ends of two novel genes were amplified by PCR, sequenced and analyzed. Results: 336 cDNA clones were analyzed and grouped into 323 species of transcript with 77 species similar to previously reported genes. Northern blot analysis identified one kidney-specific, one rarely expressed and lung-specific, and another relatively testis-specific gene. Two novel genes were cloned. One was 4.1 kb in length and encoded a 390-amino acid zinc-finger protein. Another was 2.5 kb and encoded a 474-amino acid protein of unknown function. Compared with the expression profile of a mouse renal proximal tubule cDNA library, this kidney library had a lower proportion of ribosomal genes and had a greater proportion of genes for signal transduction and DNA or RNA binding. Conclusions: Rare or novel genes could be more easily isolated from this library for molecular study of hypertension and renal pathophysiology.

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          Isolation of genes identified in mouse renal proximal tubule by comparing different gene expression profiles.

          An expression profile is a list based on a large scale sequencing of 1000 cDNA clones, showing the expressed genes and the abundance of their transcripts in a given cell or tissue (Okubo K et al: Nature Genet 2:173, 1992). We constructed an expression profile of mouse renal proximal tubules (PT) carefully isolated by microdissection in order to characterize its gene expression. Altogether 1000 clones were analyzed; there were 646 types of transcripts in PT, among which 196 were identical or homologous to the previously reported genes. The most abundant transcript was kidney-androgen regulated protein. By comparing the expression profile of PT with those obtained from other sources, several genes were identified only in PT. They included known transcripts and transcripts that were not homologous to the known genes. Three (GS4001, 3991, and 4059) of the non-homologous genes were analyzed by Northern blotting and in situ hybridization, and GS4001 and 4059 were predominantly expressed in the kidney, whereas GS3991 was detected in the liver as well as in the kidney. The sequence analysis of the full-size cDNAs demonstrated that GS4001 was a new member of aspartic proteinases and GS4059 was a novel gene. It also revealed that GS3991 was a mouse homologue of SA gene known to be expressed in PT. The expression profile of mouse PT and its comparison with those of other tissues and cells provide an alternate way of isolating genes predominantly expressed in PT, and also provides probes to study the molecular mechanisms of gene expression in the kidney.
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            Characterization of a large population of mRNAs from human testis.

            We present the results of single-pass sequencing of 779 expressed sequence tags from normal human testis cDNA clones. Of the sequences generated, 319 (41%) appeared to be completely unknown and are likely to represent new genes, and 289 (37%) were identified based on exact or nonexact matches to sequences in public databases. In analyses of hybridization of four tissues, testis, brain, liver, and kidney, 6 of 12 cDNAs clones revealed testis-specific expression. This argues for the value of the combination of random sequencing and analysis of cellular expression for large-scale characterization of gene expression in the testis.

              Author and article information

              S. Karger AG
              July 2000
              21 June 2000
              : 85
              : 3
              : 258-266
              aInstitute of Clinical Medicine, National Yang-Ming University, School of Medicine, and bDepartment of Medicine, College of Medicine, National Taiwan University, Taipei, Taiwan
              45670 Nephron 2000;85:258–266
              © 2000 S. Karger AG, Basel

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              Figures: 2, Tables: 2, References: 21, Pages: 9
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