JoVE Journal

Genetics

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Nanyang Technological University

Discrimintion and Mapping of the Primary and Processed Transcripts in Maize Mitochondrion Using a Circular RT-PCR-based Strategy

We present a circular RT-PCR-based strategy by combining circular RT-PCR, quantitative RT-PCR, RNA 5' polyphosphatase-treatment, and Northern blot. This protocol includes a normalization step to minimize the influence of unstable 5' triphosphate, and it is suitable for discriminating and mapping the primary and processed transcripts stably accumulated in maize mitochondrion.

Rozdziały w tym wideo

0:04

Title

0:39

Preparation of Crude Mitochondrion from Maize Developing Kernels

1:45

RNA 5’ Polyphosphatase Treatment and Circularization

2:33

Reverse Transcription and Normalization

3:30

PCR Amplification and Determination of Transcript Termini

4:34

Verification of the cRT-PCR Mapping Results by RNA Gel Blot Hybridization

5:00

Discrimination of Primary and Processed 5’ Ends

5:26

Determination of Maize mRNA Termini Using the Circular RT-PCR-based Strategy

7:05

Conclusion

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