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Sepiella maindroni neuropeptide Y and recombinant expression method and application thereof

A technology of the squid without needles and its expression method, which is applied in the field of neuropeptide Y and its recombinant expression in the cuttlefish without needles, can solve the problems of low feed intake rate and high dependence, so as to improve the feed intake rate and promote growth and development Effect

Inactive Publication Date: 2018-12-18
ZHEJIANG OCEAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Although this kind of feed can provide various nutritional elements for squid, it cannot solve the problem that squid is highly dependent on fresh baits such as mysis and miscellaneous fish. The feed intake of the above types of feed is still not high
However, there is no report about the neuropeptide Y of mansoni-free cuttlefish and the recombinant expression of neuropeptide Y of mansoni-needle-free squid

Method used

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  • Sepiella maindroni neuropeptide Y and recombinant expression method and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] In this embodiment, a recombinant expression method of neuropeptide Y of Manson's needleless squid comprises the following steps:

[0042] (1) Design and synthesis of a recombinant gene expressing neuropeptide Y

[0043] The applicant constructed the coding gene bank of squid mansoni, screened the coding gene of squid neuropeptide Y (as shown in SEQ ID No.7) from the gene bank, and predicted the needleless squid mansoni according to the coding gene. The full-length coded amino acid sequence of squid neuropeptide Y precursor peptide is shown in SEQ ID No. 5. This embodiment intercepts the mature short peptide of neuropeptide Y shown in SEQ ID No. 1 from the full-length coded amino acid sequence. And the enzyme-cleaved modified peptide segment (as shown in SEQ ID No.3) of the amino acid sequence C-terminus shown in SEQ ID No.5, will have the secretory signal peptide (in neuropeptide Y maturation) with the amino acid sequence shown in SEQ ID No.2 The N-terminus of the neu...

Embodiment 2

[0061] The transfected cell culture solution obtained in Example 1 was concentrated, purified by HPLC, dissolved in pure water, and freeze-dried to make a powder injection for future use.

[0062] In the period of rapid growth (weight of 20-50g) of the squid without needles, take the neuropeptide Y powder injection of the squid without needles, dissolve it in normal saline, and make a NPY solution with a concentration of 10mM, and inject it subcutaneously. Needle squid was injected once every 3 days, 50 μL each time, and continued until harvested.

[0063] During the period of subcutaneous injection of neuropeptide Y in cuttlefish, cuttlefish soft granule feed disclosed in the Chinese invention patent application with application number CN 201710798826.0 was used to feed cuttlefish at 8:00 am and 5:00 pm every day. Feed once, and the amount of each feeding is 2% of the total mass of squid mansoni.

[0064] At the same time, a control group was set up, and the control group wa...

Embodiment 3

[0069] The transfected cell culture solution obtained in Example 1 was concentrated, purified by HPLC, dissolved in pure water, and freeze-dried to make a powder injection for future use.

[0070] The needleless cuttlefish neuropeptide Y powder and the soft granule feed of the needleless cuttlefish published in the Chinese invention patent application with the application number CN 201710798826.0 were mixed evenly at a mass ratio of 1:1000 and 1:10000, and fed to Mansoni The needleless cuttlefish is fed once every day at 8 o'clock in the morning and at 5 o'clock in the afternoon, and the amount of each feeding is 2% of the total mass of the squid without needles; it is continuously fed until harvested.

[0071] At the same time, a control group was set up, and the control group was only fed the above-mentioned squid soft pellet feed without adding neuropeptide Y from squid.

[0072] The growth and development status of squid mansoni before and after feeding and the feed intake...

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Abstract

The invention discloses a sepiella maindroni neuropeptide Y and a recombinant expression method and application thereof. The method comprises: (1) acquiring a recombinant gene expressing the sepiellamaindroni neuropeptide Y, wherein the recombinant gene comprises a secretion signal peptide gene sequence, a neuropeptide Y gene sequence and an enzyme digestion-modified peptide fragment gene sequence orderly arranged from the end 5' to end 3', (2) inserting the recombinant gene into a backbone plasmid to obtain a recombinant plasmid, (3) transfecting a host cell with the recombinant plasmid to obtain a transfected cell, and (4) culturing the transfected cell and collecting the transfected cell culture liquid to obtain the sepiella maindroni neuropeptide Y. The method realizes recombinant expression and exocytosis expression of the sepiella maindroni neuropeptide Y. The recombinantly expressed neuropeptide Y is applied to sepiella maindroni through feeding or hypodermic injection so thatthe feeding rate of sepiella maindroni is significantly improved and the growth and development of sepiella maindroni are significantly promoted.

Description

technical field [0001] The invention belongs to the technical field of squid raising, and in particular relates to a squid neuropeptide Y and its recombinant expression method and application. Background technique [0002] Sepiella japonica (Sepiella japonica), commonly known as cuttlefish, belongs to the phylum Molluscs, the class Cephalopoda, the order Sepia, the family Sepia, the family Sepia, and the genus Sepia. Squid are distributed along the southeast coast of China, along the coasts of Russia, Japan, Korea, and Southeast Asia. The Zhoushan sea area is the most important distribution area, and it is a very important economic species in the world. Squid meat is delicious, unique in flavor and high in nutritional value. It is deeply loved by consumers. The dry product is called "boiled squid", and the products are sold well in Southeast Asia and coastal areas of my country. [0003] At present, the research and development of bait for artificial culture of squid manson...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/435C12N15/12C12N15/85A01K61/00A23K50/80A23K20/147
CPCA01K61/00A23K20/147A23K50/80C07K14/43504Y02A40/81
Inventor 杨静文王天明宋伊敏梁晶陈旭吕振明
Owner ZHEJIANG OCEAN UNIV
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