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JoVE Journal

Genetics

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

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07:26 min

July 29th, 2019

July 29th, 2019

6,051 Views

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

Transcrição

This protocol is used for mapping and discrimination of the primary and processed transcripts in maize mitochondria. This technique includes an RNA normalization step and it minimize the influence of unstable five prime triphosphate that hinders a

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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.

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