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'C299' dehydrin protein Dehydrin-S of cynodon dactylon and encoding gene and probe thereof

A dehydrin and probe technology, used in genetic engineering, plant genetic improvement, recombinant DNA technology, etc.

Active Publication Date: 2015-10-28
SHANGHAI JIAO TONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there is no literature report related to the structure of bermudagrass 'C299' dehydrin protein and its coding gene sequence

Method used

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  • 'C299' dehydrin protein Dehydrin-S of cynodon dactylon and encoding gene and probe thereof
  • 'C299' dehydrin protein Dehydrin-S of cynodon dactylon and encoding gene and probe thereof
  • 'C299' dehydrin protein Dehydrin-S of cynodon dactylon and encoding gene and probe thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Example 1 Cloning of Dehydrin-S Gene of Bermudagrass 'C299'

[0049] 1. Acquisition of plant material

[0050] Take bermudagrass 'C299' leaf tissue for RNA extraction;

[0051] 2. Extraction of RNA

[0052] Use "RNA prep pure plant total RNA extraction kit" to extract total RNA (Trizol: Invitrogen), use formaldehyde denaturing gel electrophoresis to identify the integrity of RNA, and then measure the purity of RNA on a spectrophotometer (Thermo Scientific NANODROP 1000 Spectrophotometer) and concentration;

[0053] 3. Full-length cloning of genes

[0054] According to the EST sequence and protein function annotation results obtained from the isolation of the Suppression Subtraction Library (SSH) established during the drought stress of Bermudagrass 'C299', the core fragment of the Dehydrin-S gene of Bermudagrass 'C299' was obtained. Using the RACE method (SMARTer TM RACE cDNA Amplification Kit: Clonetech) for full-length cDNA cloning in three stages:

[0055] (...

Embodiment 2

[0067] Example 2 Sequence Information and Homology Analysis of Dehydrin-S Gene of Bermudagrass ‘C299’

[0068] The full-length open reading frame sequence of the bermudagrass 'C299' Dehydrin-S gene of the present invention is 495 bp, and the detailed sequence is shown in SEQ ID NO.3. The amino acid sequence of Bermudagrass 'C299' Dehydrin-S protein was deduced according to the sequence of the open reading frame, with a total of 164 amino acid residues, a molecular weight of 16.7kDa, and an isoelectric point (pI) of 8.81. For the detailed sequence, see SEQ ID NO.4 the sequence shown;

[0069] The open reading frame sequence of Bermudagrass 'C299' Dehydrin-S and the amino acid sequence of its encoded protein were compiled in Non-redundant GenBank+EMBL+DDBJ+PDB and Non-redundant GenBank CDS translations+PDB+SwissProt+Superdate+ Nucleotide and protein homology searches were carried out in the PIR database, and it was found that it had 82% identity at the nucleotide level with ...

Embodiment 3

[0070] Example 3 Expression differences of bermudagrass 'C299' Dehydrin-S gene in different stages of drought stress

[0071] 1. Material acquisition: The leaves of Bermudagrass 'C299' materials in different stages of drought stress (0 day, 5 days, 10 days) were sampled. Wrap the samples with aluminum platinum paper and put them into liquid nitrogen immediately, and then transfer them to -80°C ultra-low temperature refrigerator for storage until use;

[0072] 2. Extraction of RNA: use RNA prep pure plant total RNA extraction (Trizol: Invitrogen); extract total RNA in different sample tissues of Bermudagrass ‘C299’;

[0073] 3. Determination of the integrity, purity and concentration of RNA: use ordinary agarose gel electrophoresis (gel concentration 0.5×TBE electrophoresis buffer; 150v, 15min) to check the integrity, the maximum rRNA brightness in the electrophoresis band should be 1.5-2.0 times the brightness of the second rRNA, otherwise it means that the rRNA sample is...

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Abstract

The invention relates to a 'C299' dehydrin protein Dehydrin-S of cynodon dactylon and a encoding gene and probe thereof. The protein is the protein showed in (a) or (b), (a) is a protein composed of the amino acid sequence showed in SEQ ID NO.4, (b) is a protein which is showed in SEQ ID NO.4 after replacing, lacking or adding one or more amino acids are conducted, is provided with activity of 'C299' dehydrin protein and is derived by (a). The invention further provides the nucleotide sequence of the 'C299' dehydrin protein and the probe for detecting the nucleotide sequence. According to the 'C299' dehydrin protein Dehydrin-S of the cynodon dactylon and the encoding gene and probe thereof, the purpose of improving drought resistant ability of turfgrass is achieved by using adversity-stressed physiological responses like drought resisting and the like of the 'C299' of the cynodon dactylon through the genetic engineering technology, theoretical foundation is provided for molecular breeding, and great application value is achieved.

Description

technical field [0001] The present invention is an important protective protein dehydrin protein Dehydrin-S and its encoding gene and probe in the response process of bermudagrass 'C299' drought stress, and specifically relates to a bermudagrass 'C299' dehydrin protein Dehydrin-S and its encoding Genes and Probes. Background technique [0002] Bermudagrass (Cynodon dactylon) is a very important warm-season turfgrass. It is a perennial herb of the Poaceae family. It has well-developed rhizomes and stolons. , good color and other advantages, it is widely used in sports grounds, playgrounds, parks and soil-fixing slope protection lawns at home and abroad. Therefore, it is one of the grass species with the highest application value and the widest application among warm-season turfgrasses. It enjoys the title of "the master grass species" of warm-season turfgrasses, and has great social, economic and ecological value. Bermudagrass has a very strong ability to resist drought, an...

Claims

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Application Information

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IPC IPC(8): C07K14/415C12N15/29C12Q1/68C12N15/11
CPCC07K14/415C12Q1/6895C12Q2600/13
Inventor 周鹏张荻任丽吕爱敏刘丹阳
Owner SHANGHAI JIAO TONG UNIV