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How useful are housekeeping genes? Variable expression in melanoma metastases

Kagedal, Bertil; Farnebaeck, Malin; Håkansson, Annika LU ; Gustafsson, Bertil and Hakansson, Leif (2007) In Clinical Chemistry and Laboratory Medicine 45(11). p.1481-1487
Abstract
Background: There is a certain difference in opinion regarding the optimal choice of housekeeping genes used as normalization factors in gene expression analysis. We have therefore examined the suitability of three housekeeping genes, hypoxanthine phosphoribosyl transferase, beta(2)-glucuronidase and beta(2)-microglobulin, for normalization of expression data from melanoma metastases. Methods: The expression of the three housekeeping genes was quantified by quantitative reverse transcription PCR in snap-frozen sections from 44 melanoma metastases, of which 19 were from patients treated with cisplatinum, dacarbazine and interferon alpha 2b. Results: The expression of each housekeeping gene varied considerably between the different... (More)
Background: There is a certain difference in opinion regarding the optimal choice of housekeeping genes used as normalization factors in gene expression analysis. We have therefore examined the suitability of three housekeeping genes, hypoxanthine phosphoribosyl transferase, beta(2)-glucuronidase and beta(2)-microglobulin, for normalization of expression data from melanoma metastases. Methods: The expression of the three housekeeping genes was quantified by quantitative reverse transcription PCR in snap-frozen sections from 44 melanoma metastases, of which 19 were from patients treated with cisplatinum, dacarbazine and interferon alpha 2b. Results: The expression of each housekeeping gene varied considerably between the different metastases. Histopathological examination of the tissue sections revealed variation in the amount of tumor cells in the tissue, necrosis, varying degrees of lymphocyte infiltration, and lymph node remnants. Based on this examination, 16 biopsies were omitted from further analysis because they had cracked, contained empty or necrotic areas, or were dominated by lymph node tissue. Even in sections with more than 90% tumor cells, a wide variation in the expression of the three housekeeping genes was found. The amount of lymphatic infiltrate in the tumors can have an effect on the expression of housekeeping genes in the metastases, whereas treatment did not seem to influence the expression. Conclusions: We conclude that the choice of housekeeping genes can have great impact on the normalization of specific genes in melanoma metastases. Furthermore, in the analysis of mRNA expression in tumor tissue, microscopic examination is of great importance to evaluate the integrity and cellular composition of the biopsy. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
beta(2)-microglobulin, hypoxanthine phosphoribosyl transferase, reverse transcription PCR, beta-glucuronidase, quantitative, normalization, endogenous control
in
Clinical Chemistry and Laboratory Medicine
volume
45
issue
11
pages
1481 - 1487
publisher
De Gruyter
external identifiers
  • wos:000251511500008
  • scopus:35648945416
ISSN
1434-6621
DOI
10.1515/CCLM.2007.303
language
English
LU publication?
yes
id
15801634-f98d-4f5e-891b-705d85d0bbf4 (old id 966349)
date added to LUP
2008-01-30 08:15:29
date last changed
2017-01-01 05:02:31
@article{15801634-f98d-4f5e-891b-705d85d0bbf4,
  abstract     = {Background: There is a certain difference in opinion regarding the optimal choice of housekeeping genes used as normalization factors in gene expression analysis. We have therefore examined the suitability of three housekeeping genes, hypoxanthine phosphoribosyl transferase, beta(2)-glucuronidase and beta(2)-microglobulin, for normalization of expression data from melanoma metastases. Methods: The expression of the three housekeeping genes was quantified by quantitative reverse transcription PCR in snap-frozen sections from 44 melanoma metastases, of which 19 were from patients treated with cisplatinum, dacarbazine and interferon alpha 2b. Results: The expression of each housekeeping gene varied considerably between the different metastases. Histopathological examination of the tissue sections revealed variation in the amount of tumor cells in the tissue, necrosis, varying degrees of lymphocyte infiltration, and lymph node remnants. Based on this examination, 16 biopsies were omitted from further analysis because they had cracked, contained empty or necrotic areas, or were dominated by lymph node tissue. Even in sections with more than 90% tumor cells, a wide variation in the expression of the three housekeeping genes was found. The amount of lymphatic infiltrate in the tumors can have an effect on the expression of housekeeping genes in the metastases, whereas treatment did not seem to influence the expression. Conclusions: We conclude that the choice of housekeeping genes can have great impact on the normalization of specific genes in melanoma metastases. Furthermore, in the analysis of mRNA expression in tumor tissue, microscopic examination is of great importance to evaluate the integrity and cellular composition of the biopsy.},
  author       = {Kagedal, Bertil and Farnebaeck, Malin and Håkansson, Annika and Gustafsson, Bertil and Hakansson, Leif},
  issn         = {1434-6621},
  keyword      = {beta(2)-microglobulin,hypoxanthine phosphoribosyl transferase,reverse transcription PCR,beta-glucuronidase,quantitative,normalization,endogenous control},
  language     = {eng},
  number       = {11},
  pages        = {1481--1487},
  publisher    = {De Gruyter},
  series       = {Clinical Chemistry and Laboratory Medicine},
  title        = {How useful are housekeeping genes? Variable expression in melanoma metastases},
  url          = {http://dx.doi.org/10.1515/CCLM.2007.303},
  volume       = {45},
  year         = {2007},
}