Related Experiment Video
Updated: May 11, 2026
Retrospective MicroRNA Sequencing: Complementary DNA Library Preparation Protocol Using Formalin-fixed Paraffin-embedded RNA Specimens
Published on: May 5, 2018
[Construction and sequence analysis of a normalized full-length cDNA library of Dendrobium officinale]
Min Jiang1, Jiang Wang, Guo-Song Wen
1Yunnan Agricultural University, Advantages of Chinese Medicinal Materials Standardization Planting in Yunnan Province Engineering Research Center, Kunming 650201, China.
Objective:
In order to obtain functional genes, a normalized stems cDNA library was constructed from medicinal plant Dendrobium officinale.
Method:
SMART (switching mechanism at 5' end of RNA transcript) cDNA synthesis combined with DSN (duplex-specific nuclease) normalization was applied to construct the normalized full-length cDNA library of D. officinale.
Result:
The titer of cDNA library was about 1.3 x 10(6) cfu x mL(-1) and the average insertion size was about 1.5 kb with high recombination rate (93.9%). Random selected 163 positive clones were sequenced at single side. Bio-information analysis indicated that 147 from 150 high-quality unique sequences matched corresponding homologous proteins, and they participated in various biological processes based on GO (gene ontology). There were 8 clones with complete coding sequence, which presumed to be full-length genes.
Conclusion:
These results showed preliminarily that we successfully constructed a normalized full-length cDNA library of D. officinale which could be used to screen the functional genes related to metabolic pathways of medicinal ingredients.
