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 inte...
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Nature Portfolio
2017
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oai:doaj.org-article:14c597db1a6d47d9ba9be1d91f2471492021-12-02T12:32:06ZSystematic comparison of 2A peptides for cloning multi-genes in a polycistronic vector10.1038/s41598-017-02460-22045-2322https://doaj.org/article/14c597db1a6d47d9ba9be1d91f2471492017-05-01T00:00:00Zhttps://doi.org/10.1038/s41598-017-02460-2https://doaj.org/toc/2045-2322Abstract 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.Ziqing LiuOlivia ChenJ. Blake Joseph WallMichael ZhengYang ZhouLi WangHaley Ruth VaseghiLi QianJiandong LiuNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 7, Iss 1, Pp 1-9 (2017) |
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DOAJ |
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DOAJ |
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EN |
| topic |
Medicine R Science Q |
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Medicine R Science Q Ziqing Liu Olivia Chen J. Blake Joseph Wall Michael Zheng Yang Zhou Li Wang Haley Ruth Vaseghi Li Qian Jiandong Liu Systematic comparison of 2A peptides for cloning multi-genes in a polycistronic vector |
| description |
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. |
| format |
article |
| author |
Ziqing Liu Olivia Chen J. Blake Joseph Wall Michael Zheng Yang Zhou Li Wang Haley Ruth Vaseghi Li Qian Jiandong Liu |
| author_facet |
Ziqing Liu Olivia Chen J. Blake Joseph Wall Michael Zheng Yang Zhou Li Wang Haley Ruth Vaseghi Li Qian Jiandong Liu |
| author_sort |
Ziqing Liu |
| title |
Systematic comparison of 2A peptides for cloning multi-genes in a polycistronic vector |
| title_short |
Systematic comparison of 2A peptides for cloning multi-genes in a polycistronic vector |
| title_full |
Systematic comparison of 2A peptides for cloning multi-genes in a polycistronic vector |
| title_fullStr |
Systematic comparison of 2A peptides for cloning multi-genes in a polycistronic vector |
| title_full_unstemmed |
Systematic comparison of 2A peptides for cloning multi-genes in a polycistronic vector |
| title_sort |
systematic comparison of 2a peptides for cloning multi-genes in a polycistronic vector |
| publisher |
Nature Portfolio |
| publishDate |
2017 |
| url |
https://doaj.org/article/14c597db1a6d47d9ba9be1d91f247149 |
| work_keys_str_mv |
AT ziqingliu systematiccomparisonof2apeptidesforcloningmultigenesinapolycistronicvector AT oliviachen systematiccomparisonof2apeptidesforcloningmultigenesinapolycistronicvector AT jblakejosephwall systematiccomparisonof2apeptidesforcloningmultigenesinapolycistronicvector AT michaelzheng systematiccomparisonof2apeptidesforcloningmultigenesinapolycistronicvector AT yangzhou systematiccomparisonof2apeptidesforcloningmultigenesinapolycistronicvector AT liwang systematiccomparisonof2apeptidesforcloningmultigenesinapolycistronicvector AT haleyruthvaseghi systematiccomparisonof2apeptidesforcloningmultigenesinapolycistronicvector AT liqian systematiccomparisonof2apeptidesforcloningmultigenesinapolycistronicvector AT jiandongliu systematiccomparisonof2apeptidesforcloningmultigenesinapolycistronicvector |
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1718394141665132544 |