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Updated: Sep 18, 2026
Preparation Of Neovascular Tissues from Human Glioma Tissues for Quantitative Proteomics Analysis of Tumor Angiogenesis
Published on: March 20, 2026
[Rapid construction and identification of full-length cDNA library of human glioma tissues]
Yan-yang Tu1, Ru-xiang Xu, Zhi-lin Yang
1Institute of Neuroscience, Zhujiang Hospital, First Military Medical University, Guangzhou 510282, China.
Objective:
To explore a method for rapid construction of a full-length cDNA library of human glioma tissues using switching mechanism at 5' end of RNA transcript (SMART).
Methods:
The total RNA was extracted from several samples of human glioma tissues and the mRNA was subsequently separated. Multiple mRNA samples were mixed to be used as the template for the first-strand cDNA synthesis. The CDS /3' PCR primer (containing Sfi IB site) was used in the first-strand reaction, and the SMART IV Oligo(dT) (containing Sfi A site) served as the short, extended template at the 5' end of the mRNA. With the above two primers, the primer-extension step generated full-length double-strand cDNA, which was digested by Sfi I restriction enzyme and ligated to the Sfi I A & B -digested lambdaTriplEx2 vector. The ligated vector was then packaged by lambda packaging extract for the final construction of the cDNA library.
Results:
The unamplified human glioma cDNA library consisted of 2.4x10(6) independent clones with a recombination rate of 100%. The titer of the amplified cDNA library was 4.5x10(9) pfu/ml, and the average exogenous inserts of the recombinants was 1.2 kb in length.
Conclusion:
A high-quality full-length cDNA library of human gliomas was constructed successfully, which may facilitate further study of the screening and cloning of new tumor suppressor genes and tissue-specific genes of human glioma.
Insights
A novel method rapidly constructed a high-quality, full-length complementary DNA (cDNA) library from human glioma tissues. This advancement aids in discovering new tumor suppressor and tissue-specific genes in gliomas.
Area of Science:
- Molecular Biology
- Genomics
- Cancer Research
Context:
- Human gliomas are aggressive brain tumors with complex genetic alterations.
- Understanding glioma at the molecular level is crucial for developing targeted therapies.
- Existing cDNA library construction methods can be time-consuming and yield limited full-length transcripts.
Purpose:
- To develop and validate a rapid method for constructing a full-length cDNA library from human glioma tissues.
- To utilize the Switching Mechanism at the 5' end of RNA Transcript (SMART) technology for efficient cDNA synthesis.
- To create a high-quality resource for future gene discovery in glioma.
Summary:
- Total RNA was extracted from human glioma tissues, and mRNA was isolated.
- First-strand cDNA synthesis employed a CDS/3' PCR primer and SMART IV Oligo(dT) primer, enabling full-length cDNA generation.
- The synthesized cDNA was ligated into a lambdaTriplEx2 vector, packaged, and resulted in a library with 2.4x10^6 independent clones and an average insert size of 1.2 kb.
Impact:
- Successfully constructed a high-quality, full-length human glioma cDNA library.
- Provides a valuable tool for screening and cloning novel tumor suppressor genes.
- Facilitates the identification of tissue-specific genes relevant to glioma development and progression.
