This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Hutchins, J. T. Articles by Kan-Mitchell, J. Search for Related Content PubMed PubMed Citation Articles by Hutchins, J. T. Articles by Kan-Mitchell, J.

Novel Gene Sequences Expressed by Human Melanoma Cells Identified by Molecular Subtraction¹

Departments of Microbiology [J. T. H., R. J. D., M. S. M., C. U.], Medicine [M. S. M.], and Pathology [J. K-M.], University of Southern California School of Medicine, Los Angeles, California 90033

Despite the variety of approaches used, only a limited number of tumor-associated antigens have been described for each histological type of tumor. In this report, we present a new strategy involving molecular subtraction to identify novel melanoma-associated gene sequences. Toward this end, 156 complementary DNA clones were isolated with a subtracted melanoma complementary DNA probe (melanoma minus lung carcinoma) after screening 2 x 10⁴ independent recombinants of a melanoma expression library by in situ plaque hybridization. These clones were then polymerase chain reaction amplified, screened for duplication, and categorized into 53 discrete genes. By applying poissonian distribution to the numbers of duplicate isolates, we found most of the genes to be rare messages, present at <1 copy/200 molecules of mRNA in a typical somatic cell. Messages specific for a type of tissue are usually expressed in this range. The expression of the 53 genes was further studied in human tumor cell lines and normal tissues. Partial sequence data obtained for 20 complementary DNA clones revealed 8 novel human genes. The mRNA transcripts for 5 of the novel genes were identified by Northern blot analysis. Thus, molecular subtraction appears to be applicable for the identification of novel tumor-associated sequences. Some of the potential advantages and limitations of this technology are discussed, including its application to the molecular characterization of immunogenic melanoma-associated antigens.

¹ This investigation was supported in part by USPHS Grants CA 43220, CA 36233, and GM 40146 awarded by the National Cancer Institute, National Institute of General Medical Sciences, National Institutes of Health, Department of Health and Human Services; by a grant from the Concern Foundation; by gifts from Alan Gleitsman and the Candle Foundation; and by a Research Fellowship for J. T. H. from the National Cancer Center, New York, NY.

² To whom requests for reprints should be addressed, at Department of Pathology, Norris 710, University of Southern California, 2025 Zonal Avenue, Los Angeles, CA 90033.

Received 9/19/90. Accepted 12/18/90.

This article has been cited by other articles:

				M. S. Mitchell, J. Kan-Mitchell, B. Minev, C. Edman, and R. J. Deans A Novel Melanoma Gene (MG50) Encoding the Interleukin 1 Receptor Antagonist and Six Epitopes Recognized by Human Cytolytic T Lymphocytes Cancer Res., November 1, 2000; 60(22): 6448 - 6456. [Abstract] [Full Text]