Lactococus lactis HKS2 having lactic acid activity and separating and screening method and application thereof

A Lactococcus lactis, separation and screening technology, applied in the field of strains, can solve the problems of no Lactococcus lactis Hainan Island geographical population patent, unstable purification effect, and few Lactococcus lactis, etc., to improve immunity and inhibit the effect of Significant, strong acid-producing effect

Active Publication Date: 2018-04-27
HAINAN UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] 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 on the remote environment.
At present, domestic patents on microecological preparations of lactic acid bacteria mainly focus on fermented strains for fertilizer production and food, and there are few patents on Lactococcus lactis added to breeding feed, let alone the geographical population of Lactococcus lactis in Hainan Island. patent

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  • Lactococus lactis HKS2 having lactic acid activity and separating and screening method and application thereof
  • Lactococus lactis HKS2 having lactic acid activity and separating and screening method and application thereof
  • Lactococus lactis HKS2 having lactic acid activity and separating and screening method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0053] Example 1 Isolation and screening of Lactococcus lactis HKS2.

[0054] The separation and screening method of the above-mentioned Lactococcus lactis HKS2 of the present invention mainly comprises the following steps:

[0055] (1) Sample collection

[0056] Seawater and live shrimp intestinal contents were collected from the waters of Wenchang shrimp farm in Hainan and tropical live Litopenaeus vannamei respectively, and a small amount of the collected seawater samples were taken for later use; Shred shrimp intestines, shake and mix into shrimp intestine mixture; take a small amount of mixture for later use, and the whole operation process is carried out on the ultra-clean bench;

[0057] (2) Expanded culture of sample strains

[0058] The seawater sample and shrimp intestinal mixture described in step (1) were respectively inoculated into the MRS liquid medium, and cultured in a shaker at 30°C and 180r / min for 24h;

[0059] (3) Preliminary isolation of strains

[00...

Embodiment 2

[0065] Example 2 Molecular identification of Lactococcus lactis HKS2.

[0066] 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 microscope, as shown in the scanning electron microscope figure 2 As shown, it can be preliminarily judged that the strain is Lactococcus lactis.

[0067] 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. Using 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, perform PCR amplification on the 16srRNA gene fragment of Lactococcus lactis HKS2 Amplified, cloned and sequenced. After sequencing, 1409bp fragments were obtained respectively. The accession number in GenBan...

Embodiment 3

[0068] Example 3 Acid-producing ability test of Lactococcus lactis HKS2.

[0069] Acid-producing ability test: first use the inoculation loop to pick out 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 in the petri dish, and cultured 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 set for 3 repetitions, and the average value was taken. Finally, it was determined that the diameter of the calcium-dissolving circle of Lactococcus lactis HKS2 reached 5.00-7.00mm, such as figure 1 shown by figure 1 The diameter of the calcium-dissolving circle shows that Lactococcus l...

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Abstract

The invention relates to Lactococus lactis HKS2 having lactic acid activity and a separating and screening method and application thereof. A classification name of the strain is Lactococus lactis HKS2which is already collected at the China Center for Type Culture Collection on 31st May, 2017 under a collection number of CCTCC NO:M2017296. The strain is obtained by separating a mixture of Hainan tropical aquaculture seawater and penaeus vannamei gastrointestinal tract. The Lactococus lactis HKS2 has no hemolytic activity, has high hydrophobic activity, self-coagulation rate, high resistance toartificial gastrointestinal fluid and cholate and high bacteriostatic ability. Aquaculture experiments verify that the Lactococus lactis HKS2 when serving as a feed additive to be fed to penaeus vannamei can improve organism immunity and promote growth, is stable in effect and has good application prospect.

Description

technical field [0001] The invention belongs to the field of strains, and relates to a lactococcus lactis. More specifically, the invention relates to a Hainan native lactococcus lactis HKS2 with lactic acid activity and its separation and screening method and application. 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 microecol...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12N1/02A23K10/18A23K50/80C12R1/46
CPCA23K10/18A23K50/80C12N1/02C12N1/20C12N1/205C12R2001/46
Inventor 谢珍玉文金顺龙昊章翔
Owner HAINAN UNIVERSITY
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