| Comparison of the unlabeled and labeled pre-mRNA splicing assays in vitro |
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| 作者姓名： | TIAN XU BU JING XIN HONG ZHI YAO and JIE YANG |
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| 作者单位： | TIAN XU BU，JING XIN HONG，ZHI YAO and JIE YANG Department of Immunology，Tianjin Medical University，Tianjin，P. R. China Clinical Biochemistry Lab，the General Hospital of Tianjin Medical University，Tianjin，P. R. China |
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| 摘 要： | Pre-mRNA splicing is a fundamental process required for the expression of most metazoan genes. It is carried out by the spliceosome that catalyzes the removal of non-coding intron sequences to ligate exons into mature mRNA prior to transport and translation. The purpose of our study is to explore whether the in vitro unlabeled pre-mRNA splicing assay could be performed as an alternative method of splicing reaction other than the radiolabeled one. Two different splicing methods in vitro , P labeled and unlabeled pre-mRNA as the substrates in the reaction, were investigated. The radiolabeled products were visualized by autoradiography while the unlabeled products were observed by Ethidium Bromide (EB) staining. As a result, although there are more unspecific bands in the EB staining assay than 32P labeled one, the RNA products of in vitro splicing could be observed clearly. This suggests that the unlabeled pre-mRNA splicing assay can be an optional substitution for the isotope-labeled assay.
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| 关 键 词： | pre-mRNA剪接 体外试验 同位素标记 未标记 比较研究 |
Comparison of the unlabeled and labeled pre-mRNA splicing assays in vitro |
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| TIAN XU BU,JING XIN HONG,ZHI YAO and JIE YANG.Comparison of the unlabeled and labeled pre-mRNA splicing assays in vitro[J].Chinese Journal of Microbiology and Immunology,2006,4(4):313-317. |
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| Authors: | TIAN XU BU JING XIN HONG ZHI YAO JIE YANG |
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| Institution: | [1]Department of Immunology, Tianjin Medical University, Tianjin, P. R. China [2]Clinical Biochemistry Lab, the General Hospital of Tianfin Medical University, Tianfin , P . R . China |
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| Abstract: | Pre-mRNA splicing is a fundamental process required for the expression of most metazoan genes. It is carried out by the spliceosome that catalyzes the removal of non-coding intron sequences to ligate exons into mature mRNA prior to transport and translation. The purpose of our study is to explore whether the in vitro unlabeled pre-mRNA splicing assay could be performed as an alternative method of splicing reaction other than the radiolabeled one. Two different splicing methods in vitro , P labeled and unlabeled pre-mRNA as the substrates in the reaction, were investigated. The radiolabeled products were visualized by autoradiography while the unlabeled products were observed by Ethidium Bromide (EB) staining. As a result, although there are more unspecific bands in the EB staining assay than 32P labeled one, the RNA products of in vitro splicing could be observed clearly. This suggests that the unlabeled pre-mRNA splicing assay can be an optional substitution for the isotope-labeled assay. |
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| Keywords: | Pre-mRNA splicing Spliceosome Ethidium Bromide staining Autoradiography |
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