Sugarcane flowering regulator protein ScFT-2 and encoding gene thereof

A flowering regulation and sugarcane technology, applied in genetic engineering, plant genetic improvement, plant peptides, etc.

Active Publication Date: 2017-11-21
SUGARCANE RES INST OF YUNNAN ACADEMY OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The purpose of the present invention is to solve the deficiencies of the existing artificial photoperiod regulation technology, provide the sugarcane flowering regulation protein ScFT-2 and its coding gene, bioinformatics analysis shows that the ScFT-2 gene has the PEBP euk conserved domain of the FT protein

Method used

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  • Sugarcane flowering regulator protein ScFT-2 and encoding gene thereof
  • Sugarcane flowering regulator protein ScFT-2 and encoding gene thereof
  • Sugarcane flowering regulator protein ScFT-2 and encoding gene thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0083] Example 1 Homologous Cloning of Sugarcane Flowering Regulatory Protein Gene ScFT-2

[0084] 1. Extraction of total RNA

[0085] Using sugarcane variety Yuetang 93-159 as the material, take the first fully developed mature leaf (i.e. +1 leaf, the same below) outside the tip core leaf at the mature stage (October 2015), and cut 0.1-0.2 g Middle leaves, ground to powder with liquid nitrogen, using Trans Zol TM Plant kit (ET121), extract sugarcane total RNA according to the instruction manual.

[0086] 2. Synthesis of the first strand of cDNA

[0087] Using sugarcane total RNA as a template, Trans One-Step gDNA Removal and cDNASynthesis Super MIX Reverse Transcription Kit (AE311) synthesizes the first strand of cDNA. System components: Take 1 μg of sugarcane total RNA, 1 μl of Anchored Oligo (dT) 18 Primer (0.5 μg / μl), 10 μl of 2×ES Reaction Mix, RT / RI Enzyme Mix 1μl, gDNA Remover 1μl, RNase-free Water to make up to 20μl. Incubate reverse transcription at 42°C for ...

Embodiment 2

[0098] The acquisition of embodiment 2 sugarcane flowering regulation protein gene ScFT-2 3 ' and 5 ' UTR

[0099] Search the sugarcane expression tag (expressed sequencetags, ESTs) database according to the sequence of the middle fragment of the target gene. As a result, the contig of the middle fragment of the gene and the complete 3' end sequence are obtained, while the 5' end sequence is incomplete, and it is necessary to design the 5' RACE gene-specific primers (Gene Specific Primer, referred to as GSP), use The RACE 5' / 3' Kit (Clontech Laboratories, Inc) kit was used to amplify the 5' end of the target gene, and the specific operation steps refer to the instructions of the RACE kit.

[0100] Using the total RNA of sugarcane mature leaves as a template, the first strand of cDNA with adapters was synthesized by reverse transcription using the reverse transcription adapter primers and reagents included in the SMARTer○R RACE kit.

[0101]5' cDNA first-strand synthesis

[...

Embodiment 3

[0117] Example 3 The full-length cDNA of the sugarcane flowering regulatory protein gene ScFT-2 and the acquisition of genomic DNA

[0118] Full-length cDNA and genomic DNA primers were designed according to the spliced ​​sequence, and a 963bp cDNA product (SEQ ID No.2 in the sequence table) and a 3932bp genome were obtained by PCR amplification including the start codon (ATG) and the stop codon (TGA). DNA sequence (sequence table SEQ ID No.3), PCR amplified product is detected by agarose gel electrophoresis with a mass percent concentration of 1.2%, and the target product is purified and recovered from the gel, and 4 μl of the recovered and purified PCR product is mixed with 1 μl Gently mix, react at room temperature for 5 minutes, connect with a PCR machine at 25°C for 30 minutes, put the ligation product in 50 μl Trans-T 1 competent cells (add the ligation product when the competent cells are just thawed), flick and mix well, and place on ice 25min, heat shock at 42°C for ...

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Abstract

The invention relates to sugarcane flowering regulator protein ScFT-2 and an encoding gene thereof, and belongs to the technical field of plant genetic engineering. The sugarcane flowering regulator protein ScFT-2 is composed of an amino acid sequence shown in SEQ ID No. 1, and nucleotide sequences of the encoding gene and genome DAN are shown as in SEQ ID No. 2 and SEQ ID No. 3. Total RNA and genome DNA are extracted from mature leaves of the sugarcane variety Yue Tang 93-159, and full-length cDNA and genome DNA of the ScFT-2 are acquired via the PCR (polymerase chain reaction) method in conjunction with the RACE (rapid amplification of cDNA ends) technique. The ScFT-2 gene herein acts on sugarcane stalk tips and young leaves, plays a part in sugarcane fluorescence regulation and ear formation and development, can promote sugarcane flowering, and has great practical value in studying the sugarcane flowering mechanism, promoting flowering of sugarcane parents, and researching and developing sugarcane fluorescence regulation techniques.

Description

technical field [0001] The invention relates to the technical field of genes in molecular biology, belongs to the technical field of plant genetic engineering, in particular to sugarcane flowering regulation protein ScFT-2 and its coding gene. Background technique [0002] The transition from vegetative to reproductive growth is a major event in the development of higher plants. The transformation of the reproductive process in vegetative tissues is regulated by both intrinsic and environmental factors. The shoot apical meristem (SAM) is a population of undifferentiated cells that develop into leaves and shoots during vegetative growth. Under the influence of environment and internal factors, SAM shoot apical meristem undergoes a specific change to produce flower (bud) primordium. Through the molecular biology analysis of Arabidopsis flowering induction, four flowering genes including autoregulatory pathway, gibberellin pathway, photoperiod pathway and vernalization pathwa...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/415C12N15/29C12Q1/68
CPCC07K14/415C12Q1/686C12Q1/6895C12Q2600/158C12Q2561/113C12Q2563/107C12Q2545/114
Inventor 林秀琴刘新龙李旭娟陆鑫刘洪博字秋艳毛钧吴转娣徐超华李纯佳
Owner SUGARCANE RES INST OF YUNNAN ACADEMY OF AGRI SCI
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