Molecular markers for identifying PHYB wild type and mutant of rice phytochrome gene

A phytochrome and molecular marker technology, applied in the field of molecular markers, can solve the problems of out-of-stock, increased identification cost, large amplified fragments, etc., to reduce costs and improve breeding efficiency.

Active Publication Date: 2017-01-04
SHANDONG RICE RES INST
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  • Claims
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Problems solved by technology

Although Takano et al. (2005) developed a method to identify phyB1 mutations in the article Distinct and Cooperative Functions of Phytochromes A, B, and C in the Control of Deetiolation and Flowering in Rice (The Plant Cell, Vol. However, the amplified fragment of this marker is relatively large (1Kb) and has a high GC content. It needs to be amplified with a high GC PCR buffer, and the PCR amplificat...

Method used

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  • Molecular markers for identifying PHYB wild type and mutant of rice phytochrome gene
  • Molecular markers for identifying PHYB wild type and mutant of rice phytochrome gene
  • Molecular markers for identifying PHYB wild type and mutant of rice phytochrome gene

Examples

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Embodiment 1

[0036] Example 1: Establishment of a molecular marker for identifying rice phytochrome gene PHYB wild type and mutants

[0037] (1) Primer design

[0038] It is known that the phyB1 mutant inserts a base C at 1644 bp inside the PHYB gene, thereby causing premature translation termination (Takano et al., 2005, Plant Cell, 2005, 17:3311-3325). Select 175bp sequences before and after the mutation site for comparison ( figure 1 ), according to sequence differences, practical dCAPS Finder 2.0 online design software ( http: / / helix.wustl.edu / dcaps / dcaps.html ) design primers, the primer sequences are as follows:

[0039] Upstream primer PHYBF4: 5'-TGTCACACAAAGCCCCAGCATCATGGATC-3'(Seq No.3);

[0040] Downstream primer PHYBR1: 5'-AAGGCTATCTGTGCTGAGCC-3' (Seq No.4).

[0041] (2) Theoretical analysis of amplified fragments

[0042] A 175bp fragment (sequence 1) can be amplified from wild-type rice genomic DNA; a 176bp fragment can be amplified from phyB mutant genomic DNA (seque...

Embodiment 2

[0047] Example 2: Using the molecular markers of the rice phytochrome gene PHYB wild type and mutants to analyze the PHYB genotypes of 6 rice materials

[0048] (1) Extraction of genomic DNA from rice leaves

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Abstract

The invention discloses molecular markers for identifying the PHYB wild type and mutant of a rice phytochrome gene. The nucleotide sequences of the markers related to phyB wild type site detection and phyB mutant site detection are shown as Seq No.1-2 respectively. A primer PHYBF4/PHYBR1 is used for carrying out PCR amplification, and an amplification product is subjected to BamH I digestion and electrophoretic analysis, wherein if the PCR product can not be digested by BamH I and is a 175bp DNA fragment, the PCR product is the wild type phyB, and if the PCR product can be digested by BamH I, and the digestion products are a 25bp DNA fragment and a 151bp DNA fragment, the PCR product is the mutant phyB1. The molecular markers have the advantages that amplification is easy and fast, and cost is low. The molecular markers can be used for effectively, rapidly and reliably identifying the phyB allelotype, and a powerful tool is provided for subsequent application of the phyB mutant in breeding population screening.

Description

technical field [0001] The invention relates to a molecular marker for distinguishing rice phytochrome gene PHYB wild type and mutant, and belongs to the field of plant biotechnology. Background technique [0002] Higher plants use photoreceptors such as phytochromes, cryptochromes, and phototropins to sense changes in light quality, light intensity, and photoperiod in their growth environment, and regulate their own growth and development. , to adapt to the surrounding environment as much as possible. Among them, phytochrome mainly senses red light (R) and far-red light (FR). The phytochrome gene family in rice includes three members: PHYA, PHYB and PHYC. Among them, the PHYB gene is located on the short arm of the third chromosome of rice and is a single copy. Takano et al. (2005) obtained multiple phyB mutants through gamma ray-induced mutation screening, which were named phyB1-phyB5. Among them, the phyB1 mutant inserts a base C at 1644 bp inside the PHYB gene, there...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
Inventor 郑崇珂谢先芝周晋军白波和亚男孙伟解丽霞
Owner SHANDONG RICE RES INST
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