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Molecular cloning, identification and analysis of lung squamous cell carcinoma-related genes
Chenyang Shen1, Hui-Zhao, Danli Wang
1Department of Thoracic Surgery, People's Hospital, Peking University, Beijing 100044, China. yangyang@public.fhnet.cn.net
Objective:
To clone and identify genes differentially expressed in human lung squamous cell carcinoma (LSCC).
Methods:
A subtracted cDNA library of human LSCC was constructed by using suppression subtractive hybridization (SSH) method. Through screening, the subtracted library clones, representing mRNAs that are truly differentially expressed in LSCC but not in normal lung tissues, were selected out to identify by semi-quantitative RT-PCR in 12 patients of LSCC and performed DNA sequencing. Nucleic acid homology searches were performed using the BLAST program. Partial novel genes were detected by Northern blot.
Results:
By this technique, we obtained 10 differentially expressed gene cDNA fragments of LSCC. Among them six were already known genes; two sequences were already identified but their functions were still unknown (hypothetical protein); two were novel (GenBank accession number were AF363068 and AY032661, respectively). The results from semi-quantitative RT-PCR showed that the transcription expression level of these clones including PPP1CB, caluminin, S100A2, HSNOV1, OCIA and AY032661 was down-regulated in 12 cases of LSCC, while the transcription of HSP90, ferritin, gp96 and AF363068 was up-regulated in same cases.
Conclusion:
SSH is a powerful technique of high sensitivity for the detection of differential gene expression in LSCC and an effective method to clone novel genes. Six already known genes identified by SSH technique have been already implicated in the pathogenesis of lung carcinogenesis, or they are involved in immunological defense mechanism in human body. Two hypothetical proteins probably also play an important role in lung cancer pathogenesis. The function of two novel genes in lung carcinogenesis is under research.
Insights
This study identified 10 differentially expressed genes in human lung squamous cell carcinoma (LSCC) using suppression subtractive hybridization. Two novel genes and two hypothetical proteins were discovered, offering new insights into LSCC pathogenesis.
Area of Science:
- Molecular biology
- Genomics
- Cancer research
Background:
- Lung squamous cell carcinoma (LSCC) is a major subtype of non-small cell lung cancer.
- Identifying differentially expressed genes is crucial for understanding LSCC pathogenesis and developing targeted therapies.
Observation:
- Suppression subtractive hybridization (SSH) was employed to construct a subtracted cDNA library for LSCC.
- This method effectively identified genes with altered expression in LSCC compared to normal lung tissues.
- Semi-quantitative RT-PCR and DNA sequencing were used for validation and identification of selected clones.
Findings:
- Ten differentially expressed gene cDNA fragments were identified in LSCC.
- Six known genes, two hypothetical proteins, and two novel genes (AF363068, AY032661) were discovered.
- Specific genes like PPP1CB, caluminin, S100A2, HSNOV1, OCIA, and AY032661 showed down-regulation, while HSP90, ferritin, gp96, and AF363068 were up-regulated in LSCC.
Implications:
- SSH is a sensitive and effective technique for detecting differential gene expression and cloning novel genes in LSCC.
- Several identified genes, including known and hypothetical ones, may play significant roles in lung cancer development and immune response.
- Further research is needed to elucidate the precise functions of the two novel genes in LSCC pathogenesis.