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Specific start factor of paddy rice callus and uses

A callus and promoter technology, applied in application, recombinant DNA technology, introduction of foreign genetic material using vectors, etc., can solve the problems of tedious complexity and long transformation period, and achieve high sensitivity, short period and great practical application value. Effect

Inactive Publication Date: 2009-02-04
BEIJING KAITUO DNA BIOTECH RES CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the above-mentioned elimination of the screening marker gene at the genetic level has the disadvantages of being cumbersome and complicated, and the transformation cycle is long.

Method used

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  • Specific start factor of paddy rice callus and uses
  • Specific start factor of paddy rice callus and uses
  • Specific start factor of paddy rice callus and uses

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Example 1. Isolation and identification of the rice callus-specific promoter rcsP

[0028] 1. Acquisition of the gene S126 specifically expressed in rice callus

[0029] RT-PCR was performed to detect the expression of 142 rice transcription factors in 7 tissues and organs, including rice seedling leaves, rice seedling leaves, rice roots, flowers, seeds, germinating embryos, and rice callus. Taking Actin as a reference, a gene specifically expressed in rice callus was obtained as a result, and it was named as S126 (NCBI nucleotide sequence number is: AK106306), such as figure 1 Shown (swimming lane 1 is the leaf of seedling, and swimming lane 2 is the leaf of growing seedling, and swimming lane 3 is root, and swimming lane 4 is flower, and swimming lane 5 is seed, and swimming lane 6 is the embryo that is germinating, and swimming lane 7 is rice callus tissue), The primer sequences for amplifying S126 are as follows:

[0030] Primer 1 (upstream primer): 5'-CACCAAGGATG...

Embodiment 2

[0057] Example 2, using the rice callus-specific promoter rcsP to control the expression of selection marker genes

[0058] One, the construction (see Figure 5 )

[0059] see Figure 5 (LB and RB are the left and right arms of T-DNA, respectively; RCSP: promoter rcsP; hpt: hygromycin resistance gene; bar: PPT resistance gene; attL1 and attL2: exchange arms of LR reaction on pENTR vector ; P35S: cauliflower virus 35S promoter; T35: cauliflower virus 35S terminator) to construct recombinant vectors pCAMBIA1301-RCSP-hpt and pCAMBIA1301-RCSP-bar containing rice callus-specific promoter rcsP and different resistance selection marker genes, The specific process is as follows:

[0060] 1. Gene amplification and intermediate vector construction

[0061] Primers for amplifying rcsP:

[0062] Primer 9 (upstream primer): 5'-CACCAACTGCAGAACCAAGTCTCTGTAGATTTCTGTCCCATCCGCC-3'

[0063] Primer 10 (downstream primer): 5'-CCGCTCGAGCGGCGGAATTCCGTCTCGTGCCGTTCTTGCGTTGCTCTCTC-3';

[0064] P...

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Abstract

This invention discloses the application of rice calli specific promoter in plant expression. The promoter is one of the following nucleotide sequences: (1) DNA sequence of SEQ ID No.1in the sequence list; and (2) the nucleotide sequence that can hybridize with the DNA sequence of SEQ ID No.1in highly precise conditions. The promoter can control the expression of screening marker gene in rice and subsequent screening against genetically modified rice. The method has such advantages as short screening period, high screening sensitivity and high safety.

Description

technical field [0001] The invention relates to a plant promoter and its application, in particular to a rice callus-specific promoter and its application in cultivating safe transgenic plants. Background technique [0002] Screening marker genes play an important role in plant genetic engineering. Transformed cells can be screened out from a large number of untransformed cells during the plant transgenic process by using the screenable marker gene. However, the selection of genes in transgenic plants has caused people to worry about a series of biosafety aspects, such as whether the selection marker gene encodes toxic substances and allergens, whether it is not conducive to changes in plant metabolism, and whether it will reduce the efficacy of therapeutic drugs , and whether it will migrate between similar species and pathogens. How to remove screening marker genes from transgenic plants has become a concern of people. At present, there are mainly two techniques for elim...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C12N15/63C12N15/82C12N15/09C12N15/65
Inventor 翟晨光夏勉王喜萍吴艳斌曹小近谢秀娟
Owner BEIJING KAITUO DNA BIOTECH RES CENT
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