Construction and characterization of a full-lengh cDNA library from non-fresh Giardia lamblia

Construction and characterization of a full-lengh cDNA library from non-fresh Giardia lamblia

Jun-Li Guo;Jie Jiang;Wen-Yu Zheng;Ming-Luan Li;Xi-Feng Tian;Xian-Min Feng;Yue-Hua Wang;Xiao-Hong Ju;Yue-Qiong Kong

【期刊名称】《亚太热带医药杂志（英文版）》 【年(卷),期】2012(000)012

【摘要】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 ×107 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.