Construction of cDNA libraries from microdissected benign and malignant thyroid tissue.
cDNA libraries were constructed from thyroid epithelial cells gained by laser capture microdissection for gene expression analysis of the progression of thyroid cancer. Six histologically diverse thyroid tissue specimens were used. A mean of 93 ng of total RNA was gained per tissue sample from a mean estimated number of 25,000 microdissected cells per sample. Analysis of randomly selected clones from six libraries showed an average insert size of 600 (range, 300-1500) bp. Preliminary sequencing of clones selected from the six libraries indicates a range of 46% to 62% known genes per library, 4% to 25% anonymous expressed sequence tags per library, and 15% to 43% novel expressed sequence tags per library. Thyroglobulin was found in normal thyroid epithelium and follicular thyroid adenoma, whereas calcitonin precursor transcripts were found in medullary thyroid carcinoma. We demonstrate production of high-quality cDNA libraries of microdissected tissue of the thyroid, which should prove useful for gene expression analysis of human thyroid tumors.
Kaserer
Klaus
K
Department of Clinical Pathology, University of Vienna Medical School, Vienna, Austria. k.kaserer@akh-wein.ac.at
Knezevic
Vladimir
V
Pichlhöfer
Bettina
B
Scheuba
Christian
C
Passler
Christian
C
Worth
Jennifer
J
Niederle
Bruno
B
Krizman
David
D
D000236:Adenoma / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D002116:Calcitonin / Q000378:metabolism; D002277:Carcinoma / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D017931:DNA Primers / Q000737:chemistry; D004273:DNA, Neoplasm / Q000032:analysis; D005260:Female; D020869:Gene Expression Profiling; D015723:Gene Library; D006801:Humans; D008297:Male; D008875:Middle Aged; D011498:Protein Precursors / Q000378:metabolism; D012334:RNA, Neoplasm / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013954:Thyroglobulin / Q000378:metabolism; D013964:Thyroid Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology
D017931:DNA Primers; D004273:DNA, Neoplasm; D011498:Protein Precursors; D012334:RNA, Neoplasm; D002116:Calcitonin; D013954:Thyroglobulin