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Application of Nat1 gene as selection marker in genetic transformation of oomycetes

A technology of genetic transformation and screening markers, applied in the field of genetic engineering, can solve the problem of no screening markers for oomycete genes

Pending Publication Date: 2021-05-18
SHANDONG AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] In view of the above-mentioned prior art, the purpose of the present invention is to provide the application of Nat1 gene as a screening marker in the genetic transformation of oomycetes, which solves the problem that no new screening markers are available in the existing oomycete gene anaplerosis experiments; and, the present invention The screening marker has a high screening efficiency for oomycete transformants

Method used

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  • Application of Nat1 gene as selection marker in genetic transformation of oomycetes
  • Application of Nat1 gene as selection marker in genetic transformation of oomycetes
  • Application of Nat1 gene as selection marker in genetic transformation of oomycetes

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Embodiment 1

[0044] Embodiment 1: Completion and screening of Phytophthora soybean Avh109 gene

[0045] 1. Clone the Nat1 gene and the target gene (Avh109 gene) fragment:

[0046] The following primers Nat1-F and Nat1-R were used to clone the Nat1 gene fragment, and the nucleotide sequence of the cloned Nat1 gene fragment was shown in SEQ ID NO.1.

[0047] Nat1-F: 5'-ACACAAGGGCCCCGTTTCGCATGGGTACCACTCTTGACG-3'; (SEQ ID NO.2)

[0048] Nat1-R: 5'-TTCGAACCCCAGAGTCCCGCTTAGGGGCAGGGCATGCT-3'; (SEQ ID NO.3)

[0049] Search the PsAvh109 gene from the NCBI database (http: / / www.ncbi.nlm.nih.gov / ), download its CDS and genome sequence, design primers G Avh109-F and G Avh109-R, and clone the Avh109 gene fragment, which The sequence is shown in SEQ ID NO.4.

[0050] G Avh109-F: 5'-CCatcgatATGCGTCTCCAGTATGCCG-3'; (SEQ ID NO.5)

[0051] G Avh109-R: 5'-GgaattcATCAGCGGTTTGTCGCC-3'; (SEQ ID NO.6)

[0052] 2. Construct the target gene (Avh109 gene) and Nat1 gene in the complementation vector pTOR:

[00...

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Abstract

The invention discloses an application of a Nat1 gene as a selection marker in genetic transformation of oomycetes. The invention discloses an antibiotic, namely nourseothricin as a selection marker in transformation of oomycetes. According to the invention, a coding gene Nat1 capable of causing the resistance of oomycetes to nourseothricin is used as a selection marker for screening transformants during phytophthora sojae transformation. Development of the selection marker solves the problem of single selection marker during transformation of oomycetes, and provides powerful guarantee for gene complementation, gene knockout, etc., during transformation of oomycetes.

Description

technical field [0001] The invention relates to the technical field of genetic engineering, in particular to the application of Nat1 gene as a screening marker in genetic transformation of oomycetes. Background technique [0002] Oomycetes is a kind of eukaryotic organism. At least 1800 species of oomycetes have been found so far. Most oomycetes are pathogenic bacteria of plants, animals and other organisms, which have caused huge economic losses to human beings. Phytopathogenic oomycetes mainly include Downy mildews, Phytophthora, Pythium and Albugo. Compared with fungi, oomycetes have some unique physiological and biochemical characteristics, such as: hyphae have no or few septa, vegetative bodies are diploid, and the main component of cell walls is cellulose. [0003] Phytophthora (Phytophthora) refers to a class of microorganisms belonging to the Oomycota (Oomycota), most of which are plant pathogenic bacteria. Diseases caused by Phytophthora cause devastating damage t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/80C12N15/65C12N1/15C12R1/645
CPCC12N15/80C12N15/65
Inventor 王群青谭新伟邹珅珅
Owner SHANDONG AGRICULTURAL UNIVERSITY
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