Dataset Information

Systematic comparison of 2A peptides for cloning multi-genes in a polycistronic vector.

ABSTRACT: Cloning of multiple genes in a single vector has greatly facilitated both basic and translational studies that require co-expression of multiple factors or multi-units of complex protein. Many strategies have been adopted, among which 2A "self-cleaving" peptides have garnered increased interest for their polycistronic nature, small size and high "cleavage" efficiency. However, broad application of 2?A peptides is limited by the lack of systematic comparison of different 2As alone or in combination. Here we characterized the effect of varying gene position and 2As on the expression of proteins encoded in bi-, tri-, or quad-cistronic constructs. Using direct cardiac reprogramming as an example, we further determined the effect of varied 2As on the efficiency of fluorescent cell labeling and cell fate conversion. We found that the expression of fluorophores decreased as it was moved towards the end of the construct while reprogramming was most efficient with the fluorophore at the second position. Moreover, quad-cistronic TPE2A constructs resulted in more efficient reprogramming than 3P2A or PTE2A constructs. We expect that the bi-, tri-, and quad-cistronic vectors constructed here and our results on protein expression ratios from different 2A constructs could serve to guide future utilization of 2A peptides in basic research and clinical applications.

SUBMITTER: Liu Z

PROVIDER: S-EPMC5438344 | biostudies-literature | 2017 May

REPOSITORIES: biostudies-literature

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Systematic comparison of 2A peptides for cloning multi-genes in a polycistronic vector.

Liu Ziqing Z Chen Olivia O Wall J Blake Joseph JBJ Zheng Michael M Zhou Yang Y Wang Li L Vaseghi Haley Ruth HR Qian Li L Liu Jiandong J

Scientific reports 20170519 1

Cloning of multiple genes in a single vector has greatly facilitated both basic and translational studies that require co-expression of multiple factors or multi-units of complex protein. Many strategies have been adopted, among which 2A "self-cleaving" peptides have garnered increased interest for their polycistronic nature, small size and high "cleavage" efficiency. However, broad application of 2 A peptides is limited by the lack of systematic comparison of different 2As alone or in combinati ...[more]

PMID: 28526819

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Project description:BackgroundIn recent years, the important functional roles of RNAs in biological processes have been repeatedly demonstrated. Computing the similarity between two RNAs contributes to better understanding the functional relationship between them. But due to the long-range correlations of RNA, many efficient methods of detecting protein similarity do not work well. In order to comprehensively understand the RNA's function, the better similarity measure among RNAs should be designed to consider their structure features (base pairs). Current methods for RNA comparison could be generally classified into alignment-based and alignment-free.ResultsIn this paper, we propose a novel wavelet-based method based on RNA triple vector curve representation, named multi-scale RNA comparison. Firstly, we designed a novel numerical representation of RNA secondary structure termed as RNA triple vectors curve (TV-Curve). Secondly, we constructed a new similarity metric based on the wavelet decomposition of the TV-Curve of RNA. Finally we also applied our algorithm to the classification of non-coding RNA and RNA mutation analysis. Furthermore, we compared the results to the two well-known RNA comparison tools: RNAdistance and RNApdist. The results in this paper show the potentials of our method in RNA classification and RNA mutation analysis.ConclusionWe provide a better visualization and analysis tool named TV-Curve of RNA, especially for long RNA, which can characterize both sequence and structure features. Additionally, based on TV-Curve representation of RNAs, a multi-scale similarity measure for RNA comparison is proposed, which can capture the local and global difference between the information of sequence and structure of RNAs. Compared with the well-known RNA comparison approaches, the proposed method is validated to be outstanding and effective in terms of non-coding RNA classification and RNA mutation analysis. From the numerical experiments, our proposed method can capture more efficient and subtle relationship of RNAs.

Dataset Information

Systematic comparison of 2A peptides for cloning multi-genes in a polycistronic vector.

Publications

Systematic comparison of 2A peptides for cloning multi-genes in a polycistronic vector.

Similar Datasets

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