A kind of Paenibacillus neutral phytase

A spore-like and phytase technology, applied in the fields of hydrolytic enzymes, plant genetic improvement, botany equipment and methods, etc., can solve the problems that cannot fully meet the requirements of feed processing, and the expression level of natural materials is low

Active Publication Date: 2011-12-21
内蒙古蒙草土壤科技有限责任公司 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] At present, the main problem of phytase in industrial production is that the expression level in natural materials is low, which cannot fully meet the requirements of feed processing.

Method used

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  • A kind of Paenibacillus neutral phytase
  • A kind of Paenibacillus neutral phytase
  • A kind of Paenibacillus neutral phytase

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Embodiment 1 produces the screening of the Paenibacillus of phytase

[0033] Soil was obtained from high-temperature irradiation environment in Xinjiang for microbial enrichment culture. Weigh 1g of soil and put it into the enrichment medium, culture at 37°C, 200r / min, for 6 days. Draw a small amount of culture solution every day for serial dilution (10 -2 ~10 -10 ), spread on the screening solid medium containing sodium phytate, and culture at a constant temperature of 37°C for 3 days. After the colonies grew out, they were repeatedly streaked and separated, and the colonies that could hydrolyze sodium phytate and have larger hydrolysis circles were selected.

Embodiment 2

[0034] Example 2 Extraction and Identification of Paenibacillus sp. Genome

[0035] With the bacterium colony in the culture medium selected in embodiment 1, pick single bacterium colony and put into LB liquid culture medium, take bacterium liquid and be used for the extraction of the genomic DNA of this bacterium. Using the bacterial genomic DNA extraction kit of Omega Company, the genomic DNA of the bacteria was extracted according to the operating instructions of the kit provided by the company. In order to identify the genus, the 16s rRNA gene sequence of the strain was amplified by polymerase chain reaction (PCR) using genomic DNA as a template.

[0036] Design primers for bacterial 16s rRNA gene sequence: F 27: 5′-AGAGTTTGATCATGGCTCAG-3′

[0037] R 1492: 5′-TACGGTTACCTTGTTACGACTT-3′

[0038] The length of the PCR product is about 1.5kb. After sequencing, it is compared with the DNA sequence in the GenBank database. It is found that the sequence is 98% similar to the 16...

Embodiment 3

[0039] The cloning of phytase gene in the Paenibacillus genome of embodiment 3

[0040] Find other Paenibacillus phytase gene sequences from Genbank, and design degenerate primers, as follows:

[0041] phyP F(EcoR I): 5′- GAATTC TTGGTTAAGTTGGAAAACGG-3′

[0042] phyP R(Not I): 5′- GCGGCCGC TCTTGGAGACAACTTTCTTG-3′

[0043] The underlined part is the recognition site of EcoR I (upstream primer F) and Not I (downstream primer R).

[0044]After the PCR product was subjected to 0.8% agarose gel electrophoresis, the observed length was about 1.3kb. T4 DNA ligase was used to connect to the T-vector (pGEM-T vector, Promega Company). After sequencing, the deduced amino acid sequence is shown as SEQ ID NO:2.

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Abstract

The invention obtains a neutral phytase gene from Paenibacillus and has a nucleotide sequence disclosed in SEQ ID NO:1. The gene can be used for producing the natural phytase with higher enzymatic activity.

Description

technical field [0001] The invention relates to a Paenibacillus neutral phytase, in particular to a gene sequence and an amino acid sequence encoding the neutral phytase. Background technique [0002] Phytase, also known as phytase, is a general term for a class of enzymes that catalyze the hydrolysis of phytic acid and phytate into inositol and phosphoric acid (or salt), and can hydrolyze phytic acid to finally release inorganic phosphorus. In recent years, it has been widely used to promote the growth and development of monogastric animals and improve the utilization rate of phosphorus in feed. Its research is of great significance to improve animal husbandry production efficiency and reduce environmental pollution; the application of phytase in food can improve the concentration of iron and zinc. Absorption is conducive to improving the nutrition of the crowd. [0003] At present, the main problem of phytase in industrial production is that the expression level in natura...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/55C12N9/16C12N15/63C12N15/81C12R1/01C12R1/84
Inventor 平淑珍赵仲麟燕永亮陆伟林敏
Owner 内蒙古蒙草土壤科技有限责任公司
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