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Strawberry auxin synthetic rate-limiting enzyme gene FaYUC11 and application

A rate-limiting enzyme and auxin technology, applied in the field of plant biology, can solve the problems of low ratio and low average yield

Active Publication Date: 2015-03-04
SHANGHAI ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] my country's strawberry production can be called a "big strawberry country" in terms of total area and total output, but there is still a certain gap compared with the "world's strawberry powerhouses" such as the United States and Spain. Very low, and significantly lower average yields

Method used

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  • Strawberry auxin synthetic rate-limiting enzyme gene FaYUC11 and application
  • Strawberry auxin synthetic rate-limiting enzyme gene FaYUC11 and application
  • Strawberry auxin synthetic rate-limiting enzyme gene FaYUC11 and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Example 1 Isolation of FaYUC11 gene and construction of virus-induced gene silencing vector

[0044] Based on the sequence information of the whole family of YUC genes in diploid forest strawberry (Liu et al. 2014), the dynamic expression patterns of the whole family genes were resolved in the octoploid cultivated strawberry fruit (receptacle and achene). It was found that FaYUC11 was the only highly expressed member of this family in achenes, and its dynamic expression pattern was consistent with that of auxin accumulation in achenes.

[0045] Design PCR amplification primers according to the sequence of the forest strawberry homologous gene, forward primer OF:

[0046] 5'-AAAATGGAGAACAATGTGTTTGGGA-3'; reverse primer OR:

[0047] 5'-GGACTAGACCTCTCTTGCAGCAT-3', as shown in SEQ ID No.3 and 4, respectively.

[0048] Using the wild-type Jiuxiang Strawberry fruit cDNA as a template, the full-type gold high-fidelity enzyme (from Beijing TransGene Company) was used for PCR ...

Embodiment 2

[0053] Embodiment 2 The acquisition of virus-induced gene silencing strawberry

[0054] The pBSK-in-dYUC11 recombinant plasmid was further digested with KpnI enzyme and SacI enzyme, and the bidirectional hairpin structure of the FaYUC11 gene fragment was cloned into the CP protein behind the 2×35S promoter of tobacco rattle virus pTRV2 (Liu et al. 2002). Downstream, the pTRV2-dYUC11 recombinant vector was obtained. Agrobacteria containing RNA1 and RNA2 of tobacco rattle virus were mixed.

[0055]Inoculate GV3101 Agrobacterium containing target gene virus-induced silencing expression vector and empty pTRV1 and pTRV2 vectors in 5mL YEP medium containing 50 μg / mL Kan, 10 μg / mL Rif and 50 μg / mL Gen and culture overnight at 28°C, and the next day follow Inoculate in induction medium (YEP, containing 10mM / L MES, 20μM / L AS, 50μg / mL Kan, 10μg / mLRif and 50μg / mL Gen) at a ratio of 1:50, shake the bacteria at 28°C until logarithmic phase (OD Value 0.6-0.8), 5000r / min centrifuge for 10m...

Embodiment 3

[0056] Example 3 Identification of virus-induced gene silencing strawberry

[0057] In order to detect whether tobacco rattle virus successfully infects strawberry, the total RNA of strawberry fruit was extracted from wild-type and injected virus original vector and recombinant FaYUC11 virus-induced silencing vector, and cDNA was synthesized by reverse transcription using random primers.

[0058] According to Jia et al. 2011, specific primers for tobacco rattle virus RNA2 were provided for RT-PCR reaction to identify virus-infected strawberry fruits. PCR product is carried out electrophoresis detection, the result finds, the strawberry fruit that the recombinant virus vector of virus original vector and FaYUC11 infects can amplify virus-specific band (such as figure 2 ), but there was no band for this purpose in wild-type strawberry. It was preliminarily determined that tobacco rattle virus had infected strawberry fruit by microinjection method.

[0059] Further, a new pair...

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Abstract

The invention relates to an agricultural biotechnology, discloses a strawberry auxin synthetic rate-limiting enzyme gene FaYUC11 and application in adjustment of fruit sizes, and provides a nucleotide sequence and a protein sequence of the gene. A method for using the FaYUC11 to adjust the fruit sizes comprises the following steps of: constructing a virus induction silent vector of the FaYUC11 gene; and transferring the silent vector in agrobacterium, infecting young green fruits of strawberries by using a micro-injection method, and establishing a transgenosis strawberry strain silenced by the virus induction FaYUC11 gene. The content of free auxin in achenes (seeds) can be reduced through gene silencing; the growth rate of fruits in vertical and horizontal diameters is reduced; simultaneously, the hardness of fruits and the content of soluble solids are influenced; and thus, the gene disclosed by the invention has good application prospect in the aspect of adjusting and controlling the fruit sizes of strawberries, and has important significance on development of functional SNP (Single Nucleotide Polymorphism) molecular markers and breeding of high-quality large strawberries.

Description

technical field [0001] The invention relates to the field of plant biotechnology, in particular to an octoploid strawberry auxin rate-limiting enzyme FaYUC11 gene and its application, and a method for using the gene to regulate the size of strawberry fruit. Background technique [0002] Strawberry (Fragaria×ananassa) belongs to the Rosaceae Fragaria plant. It is one of the important fresh fruit crops and a model species for the study of important functional genes of Rosaceae plants. The genetic and molecular biology studies related to fruit development are becoming more and more important. Having attention. Among them, the regulation of phytohormones is the key to fruit development and quality formation. [0003] Strawberry fruit belongs to the "false fruit", which develops from the receptacle, and the botanical fruit is the seeds dotted on its surface, which are also called achenes. As early as the middle of last century, Nitsch found that the growth and development of st...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/53C12N9/02C12N15/84C12N15/66A01H5/00
Inventor 段可谢为发高清华张玲
Owner SHANGHAI ACAD OF AGRI SCI
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