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Published on: July 15, 2018
Construction and characterization of a full-lengh cDNA library from non-fresh Giardia lamblia
Jun-Li Guo1, Jie Jiang, Wen-Yu Zheng
1School of Laboratory Medicine, Jilin Medical College, Jilin 132013, PR China.
Objective:
To construct rapidly a full-length cDNA library from nanogram amounts total RNA of Giardia lamblia (G. lamblia) trophozoites stocked in RNA stabilization reagent.
Methods:
Total RNA of Giardia was extracted using Trizol reagent. A full-length cDNA library of G. lamblia trophozoites was constructed by a long-distance PCR (LD-PCR) method. The recombinant rate and the coverage rate of full-length clones of the library were evaluated. The inserted fragments were identified and sequenced by PCR amplification.
Results:
The titer of cDNA library was 3.85 × 10(7) pfu/mL. The length of inserted fragments ranged from 0.4 to 2.5 kb, and the recombination efficiency accounted for 100% (20/20). The coverage rate of full-length clones is high (17/20).
Conclusions:
The RNA stabilization reagent may be used to fix the cells and prevent the RNA in cells even though delivered under normal atmospheric temperature. The long-distance PCR can be used to construct a full-length cDNA library rapidly and it needs less RNA than the traditional method from mRNA.
