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A kind of enterococcus faecalis hkf7 with lactic acid activity and its screening culture method and application

A technology of enterococcus faecalis and activity, applied in the field of strains, can solve the problems of no geographical population of Lactococcus lactis and enterococcus faecalis in Hainan Island, unstable purification effect, poor purification effect, etc., to improve the body's immunity, inhibit Remarkable effect, strong acid production effect

Active Publication Date: 2020-12-01
HAINAN UNIVERSITY
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  • Abstract
  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

The strains of Bacillus licheniformis in different growth environments have a good effect on the purification of aquaculture water in the local environment, but they often show poor purification effect and unstable purification effect in the remote environment.
[0006] At present, the domestic patents on lactic acid bacteria microecological preparations mainly focus on fermented strains for fertilizer production and food. There are few patents on Enterococcus faecalis added to breeding feed, and there are no patents on Lactococcus lactis and Enterococcus faecalis. related patents

Method used

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  • A kind of enterococcus faecalis hkf7 with lactic acid activity and its screening culture method and application
  • A kind of enterococcus faecalis hkf7 with lactic acid activity and its screening culture method and application
  • A kind of enterococcus faecalis hkf7 with lactic acid activity and its screening culture method and application

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] Embodiment 1 The isolation and screening method of Enterococcus faecalis HKF7 mainly comprises the following steps:

[0051] (1) Sample collection

[0052] Seawater samples were collected from the waters of Wenchang Shrimp Farm in Hainan, and a small amount of the collected samples were used for later use. The entire operation process was carried out on a purification ultra-clean bench;

[0053] (2) Expanded culture of sample strains

[0054] Take 0.1mL of the seawater sample collected in step (1) and inoculate it into a centrifuge tube (10mL / 50mL, V / V) filled with MRS liquid medium, one tube for each sample, at 30°C, 180r / min, shaker medium shaking culture for 24 hours;

[0055] (3) Preliminary isolation of strains

[0056] The separation and purification of the bacterial strains were combined with the plate streaking method and the coating method. For the bacterial strains cultivated in step (2), 0.1 mL of the bacterial liquid was applied to the MRS solid medium (c...

Embodiment 2

[0062] Example 2 Molecular identification of Enterococcus faecalis HKF7.

[0063] In order to identify the strains, the cell morphology and some physiological and biochemical reactions of the tested strains were first observed by scanning electron microscopy, as shown in the micrograph figure 2 As shown, it can be preliminarily judged that the strain is Enterococcus faecalis.

[0064] Use the bacterial universal primers to carry out PCR identification on the screened bacterial strains: extract the template DNA according to the operating instructions of the bacterial DNA extraction kit. Use the upstream primer 5'-AGAGTTTGATCCTGGCTCA-3' of the 16S rRNA conservative sequence, see SEQ ID No.2 and the downstream primer 5'-GGTTACCTTGTTACGACTT-3, see SEQ ID No.3, to perform PCR amplification on the 16srRNA gene fragment of Enterococcus faecalis HKF7 Amplified, cloned and sequenced. After sequencing, 1422bp fragments were obtained respectively. The accession number in GenBank was MF...

Embodiment 3

[0065] Example 3 Acid-producing ability test of Enterococcus faecalis HKF7.

[0066] Acid production ability test: first use the inoculation loop to pick the target strain of primary screening, inoculate the strain in the MRS liquid medium aseptically, and then shake and culture it in a 30°C incubator at 180rpm for 24 hours, and set aside; prepare the MRS solid medium ( Add 1% light CaCO 3 ), pour 20mL culture medium into each petri dish, and use diluted 10 8 100 μL of cfu / mL bacterial solution was spread on the solid medium, and the culture dish was placed in a 30°C incubator for 48 hours, and the diameter of the calcium dissolution circle was measured with a vernier caliper. Each sample was repeated three times, and the average value was taken. Finally, it was determined that the diameter of the calcium-dissolving circle of Enterococcus faecalis HKF7 was 5.00-7.00mm, such as figure 1 shown by figure 1 The diameter of the calcium-dissolving circle shows that Enterococcus f...

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Abstract

The invention relates to an Enterococcus faecalis HKF7 with lactic acid activity and a screening and culturing method and application thereof. The bacterial strain of the present invention is classified as: (Enterococcus faecalis) HKF7, and has been preserved in the China Center for Type Culture Collection, with the preservation number CCTCC NO: M2017297, and the preservation date is May 31, 2017. The strain was isolated and screened from tropical cultured seawater in Hainan and the intestinal tract of live Litopenaeus vannamei in tropical Hainan. The Enterococcus faecalis HKF7 of the present invention has no hemolytic activity, good hydrophobic activity, high self-aggregation rate, artificial gastrointestinal fluid tolerance, bile salt tolerance, and strong antibacterial ability. The Enterococcus faecalis HKF7 of the present invention is used as Feeding Litopenaeus vannamei as a feed additive has been verified through breeding experiments that it can better improve the body's immunity and promote the growth of prawns, and the effect is stable. It is a strain with good application prospects.

Description

technical field [0001] The invention belongs to the field of strains, and relates to an Enterococcus faecalis. More specifically, the invention relates to an Enterococcus faecalis HKF7 with lactic acid activity, a screening culture method and application thereof. Background technique [0002] In order to pursue high-efficiency and rapid breeding benefits, the breeding density has been continuously increased and the scale of farming has been expanded, resulting in a decrease in the growth rate of aquatic animals, an increase in the bait coefficient, frequent occurrence of common diseases such as vibriosis, and the abuse of chemical fishery drugs such as antibiotics. This not only increases the cost of farming, but also poses a huge threat to the quality and safety of aquatic products and human health. Therefore, finding an efficient, healthy and safe breeding method has become a research focus of current aquaculture technology. [0003] The application of microecological pre...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12N1/02A23K50/80A23K10/18C12R1/01
CPCA23K10/18A23K50/80C12N1/02C12N1/20C12R2001/46C12N1/205Y02A40/818
Inventor 谢珍玉文金顺章翔龙昊
Owner HAINAN UNIVERSITY
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