Subtraction hybridization cDNA libraries from colon carcinoma and hepatic cancer

Schweinfest, Clifford W.; Henderson, Kelly W.; Gu, Jianren; Kottaridis, S. D.; Besbeas, S; Panotopoulou, Evi; Papas, Takis S.

doi:10.1016/0735-0651(90)90042-e

Cited by 68 publications

(38 citation statements)

References 8 publications

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“…After poly(A) + RNA isolation (Chomczynski and Sacchi, 1987), day 10, 11 and 12 liver cDNA libraries were constructed (Gubler and Ho man, 1983;Mishra et al, 1998). Subsequently, subtracted libraries were prepared by conventional techniques (Schweinfest et al, 1990). We obtained and partially sequenced a total of 64 clones.…”

Section: Methodsmentioning

confidence: 99%

Elf3 encodes a novel 200-kD β-spectrin: role in liver development

et al. 1999

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b-spectrins are crucial for the maintenance of cell shape, the establishment of cell polarity, and the formation of distinct membrane domains. Our strategy for identifying genes important for hepatocyte polarity has been to utilize subtractive hybridization of early embryonic mouse cDNA liver libraries. As a result, we have cloned three isoforms of a novel b-spectrin elf (embryonic liver b-fodrin), and here we report the analysis of elf3, the longest isoform (8172 nt). ELF3 comprises 2154 residues with an overall similarity of 89.0% and 95.3% to mouse b-spectrin (bSpIIS1) at the nucleotide and amino acid level, respectively. ELF3 is characterized by an actinbinding domain, a long repeat domain, and a short regulatory domain remarkable for the absence of a PH domain. Linkage analysis reveals that elf3 maps to mouse chromosome 11 between D11Bir6 and D11Xrf477, a di erent chromosomal locus from that of the other four spectrin genes. Northern blot analysis utilizing an elf3 3'-UTR probe demonstrates an abundant 9.0-kb transcript in brain, liver, and heart tissues. Western blot with a polyclonal antibody against ELF identi®es a 200 kD protein in mouse liver, brain, kidney, and heart tissues. Immunohistochemical studies demonstrate ELF labeling of the basolateral or sinusoidal membranes surface as well as a granular cytoplasmic pattern in hepatocytes. Antisense studies utilizing cultured liver explants show a vital role of elf3 in hepatocyte di erentiation and intrahepatic bile duct formation. The di erential expression, tissue localization, and functional studies demonstrate the importance of elf3 in modulating interactions between various components of the cytoskeleton proteins controlling liver and bile duct development.

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Section: Methodsmentioning

confidence: 99%

Elf3 encodes a novel 200-kD β-spectrin: role in liver development

et al. 1999

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“…The cloning of cDNAs that encode proteins specific to hair cells, supporting cells, and other cells of the inner ear is, in turn, retarded by the limited numbers of these cells in an animal. This disadvantage potentially can be circumvented through enrichment of specific cDNAs by subtractive hybridization (1)(2)(3) and library normalization (4). That only a few inner ear-specific cDNAs have been obtained from libraries constructed by this approach (2,3), however, suggests that the efficient cloning of cDNAs specific to sensoryepithelial cell types requires libraries constructed from more abundant sources than whole cochleae.…”

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confidence: 99%

Molecular markers for cell types of the inner ear and candidate genes for hearing disorders

Heller

Sheane

Javed

et al. 1998

Proc. Natl. Acad. Sci. U.S.A.

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To identify genes expressed in the vertebrate inner ear, we have established an assay that allows rapid analysis of the differential expression pattern of mRNAs derived from an auditory epithelium-specific cDNA library. We performed subtractive hybridization to create an enriched probe, which then was used to screen the cDNA library. After digoxigenin-labeled antisense cRNAs had been transcribed from hybridization-positive clones, we conducted in situ hybridization on slides bearing cryosections of late embryonic chicken heads, bodies, and cochleae. One hundred and twenty of the 196 clones analyzed encode 12 proteins whose mRNAs are specifically or highly expressed in the chicken's inner ear; the remainder encode proteins that occur more widely. We identified proteins that have been described previously as expressed in the inner ear, such as ␤-tectorin, calbindin, and type II collagen. A second group of proteins abundant in the inner ear includes five additional types of collagens. A third group, including Coch-5B2 and an ear-specific connexin, comprises proteins whose human equivalents are candidates to account for hearing disorders. This group also includes proteins expressed in two unique cell types of the inner ear, homogene cells and cells of the tegmentum vasculosum.Developmental analysis of the inner ear is hindered by the paucity of molecular markers for specific cell types. At present, most cells in the cochlea and vestibular organs cannot be unambiguously identified before they display their characteristic mature forms. The cloning of cDNAs that encode proteins specific to hair cells, supporting cells, and other cells of the inner ear is, in turn, retarded by the limited numbers of these cells in an animal. This disadvantage potentially can be circumvented through enrichment of specific cDNAs by subtractive hybridization (1-3) and library normalization (4). That only a few inner ear-specific cDNAs have been obtained from libraries constructed by this approach (2, 3), however, suggests that the efficient cloning of cDNAs specific to sensoryepithelial cell types requires libraries constructed from more abundant sources than whole cochleae. To address this problem, we constructed a cDNA library from sensory epithelia of the chicken's cochlea and developed an enriched probe with which to identify cDNAs encoding proteins expressed by specific cell types of the inner ear.The identification of proteins specific to the internal ear is also potentially useful in the study of heritable human deafness, which afflicts about one individual in a thousand. Several proteins that have been identified as markers for specific aural cell types recently have been advanced as candidate genes for human hearing disorders (for reviews see refs. 5-8). Some of the genes identified by the strategy described in this paper are likely to have human equivalents that cause hearing disorders when mutated. MATERIALS AND METHODSLibrary Construction and Screening. Basilar papillae from 250 late-embryonic (E, embryonic day) (8% E14...

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“…cDNA's were prepared from the`normal' breast cell line Hs578Bst and the tumor cell line Hs578T as described (Schweinfest et al, 1990;Salesiotis et al, 1995;Burger et al, 1996).…”

Section: Subtractive Cloningmentioning

confidence: 99%

“…In order to identify and isolate genes whose expression is associated with breast carcinomas in various stages of neoplastic development, we employed subtractive-hybridization cloning and di erential display-PCR (Schweinfest et al, 1990;Liang and Pardee, 1992;Salesiotis et al, 1995;Burger et al, 1996). RNA was prepared from two breast cell-lines derived from a tumor and the adjacent normal tissue of the same patient.…”

Section: Introductionmentioning

confidence: 99%

Breast cancer genome anatomy: correlation of morphological changes in breast carcinomas with expression of the novel gene product Di12

Burger

Zhang

et al. 1998

Oncogene

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To determine which genes may be activated or inactivated during breast cancer development, we employed two cloning strategies (subtractive hybridization and di erential display) using RNA samples from a human breast tumor and its matching normal breast cell line. Of 950 clones isolated, 102 cDNA inserts were analysed by DNA sequencing and database searching. We found 30 clones that were obviously unidenti®ed, with no signi®cant homology to any listed human gene. We focused upon one of the novel genes, Di12, that is di erentially expressed as a 1.35 kb RNA in breast cancer tissues and cell-lines, and in several normal tissues. A full length cDNA of this gene was cloned, and its DNA sequence revealed an open reading frame of 339 amino acids. Antibodies to the ten N-terminal amino acids were developed to investigate the expression of Di12 in breast cancer cell-lines and tumors. The Di12 protein was found in tissue sections of in®ltrating ductal carcinomas (IDCs), but not in benign or normal breast specimens. RT ± PCR analysis con®rmed expression of Di12 in 80% of in®ltrating ductal carcinomas (IDCs). As IDC constitutes *70% of breast cancers seen clinically, the level of Di12 expression may be predictive of disease progression.

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Subtraction hybridization cDNA libraries from colon carcinoma and hepatic cancer

Cited by 68 publications

References 8 publications

Elf3 encodes a novel 200-kD β-spectrin: role in liver development

Elf3 encodes a novel 200-kD β-spectrin: role in liver development

Molecular markers for cell types of the inner ear and candidate genes for hearing disorders

Breast cancer genome anatomy: correlation of morphological changes in breast carcinomas with expression of the novel gene product Di12

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