Humanized pika leptin protein, gene coding same and application thereof
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A leptin protein, humanized technology, applied in the field of genetic engineering, can solve problems such as being in the blank
Inactive Publication Date: 2014-07-09
CHINA ACAD OF SCI NORTHWEST HIGHLAND BIOLOGY INST
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Problems solved by technology
However, most of the leptin protein products developed and utilized at home and abroad are isolated from human bodies living in ordinary environments, while leptin proteins isolated from special cold-adapted animals under cold environmental pressure are rarely reported. Functional research not only helps people understand the mechanism of animals' ecological adaptation to extreme environments, but also provides a scientific basis for the development and utilization of their products. Research in this area is currently blank at home and abroad.
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Embodiment 1
[0048] Example 1 Artificially synthesized humanized pika leptin gene sequence
[0049] The sequence of SEQ ID No.1 was synthesized by artificial gene synthesis, and the gene synthesis was completed by Shanghai Sangon Bioengineering Co., Ltd.
Embodiment 2
[0050] Embodiment 2 Preparation of recombinant humanized pika leptin protein
[0051] 1. Experimental materials and solution preparation
[0052] (1) Expression vector and engineering bacteria
[0053] The expression vector used to express the recombinant pika leptin protein is pMAL-c2x (NewEngland Biolabs, NEB Company), and the engineering bacteria is Escherichia coli BL21 (DE3).
[0054] (2) Kits, tool enzymes and biochemical reagents
[0055] Restriction enzymes and Taq enzymes are products of Dalian Bao Bioengineering Co., Ltd., blood biochemical detection kits were purchased from Zhongsheng Beikong Biotechnology Co., Ltd., yeast extract (Yeast extract) and tryptone (Tryptone) were purchased from Oxford Co. Products, other antibiotics and biochemical reagents are products of sigma company.
[0056] (3) Protein purification chromatography column and its medium
[0057] All chromatographic media and their chromatographic columns are products of GE Healthcare, USA. The N...
Embodiment 3
[0090] Example 3 Application of humanized pika leptin protein in improving blood sugar and blood lipids
[0091] 1. Establishment of mouse model of hyperglycemia and hyperlipidemia
[0092] (1) Experimental animals
[0093] 40 male weaned (21 days) mice weighing about 25 g (purchased from the Experimental Animal Center of Hebei Medical University). Single-cage feeding, room temperature 18-25°C, relative humidity 50%-80%. 12 hours of light per day, free access to food and water. After 3 days of adaptation, blood was collected through the inner canthus, serum was separated, and baseline biochemical indicators were measured.
[0094] (2) High-fat feed formula: make containing lard 28%, white sugar 30%, milk powder 1%, egg yolk 18%, the high-fat feed of basic feed 33%.
[0095] (3) Establishment process of high-fat and high-glucose animal model
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Abstract
The invention provides a humanized pika leptin protein, a gene coding the protein. The nucleotide sequence and the amino acid sequence are respectively shown as SEQ ID No. 1 and SEQ ID No. 2. The invention further provides an expression and purification method of the humanized pika leptin recombinant protein which is soluble. The invention builds an animal model of hyperglycemia and hyperlipidemia, and uses the humanized pika leptin recombinant protein and a human leptin protein to cure, and results show that the humanized pika leptin protein, similar with the human leptin protein, can obviously reduce blood glucose concentration and total cholesterol, triglyceride levels of serum, reduce content of visceral white adipose tissue and accumulation of intracellular fat, and promote expression level of key genes participating glycometabolism and lipid metabolism in liver. Compared with human recombinant leptin protein, the humanized pika leptin protein has a relatively obvious effect for improving blood glucose and reducing blood lipid.
Description
technical field [0001] The invention belongs to the field of genetic engineering, and in particular relates to a humanized pika leptin protein, a gene encoding the protein and an application thereof. Background technique [0002] The unique animal and plant resources of the Qinghai-Tibet Plateau are unique in the world. The extreme pressure environment of high cold, low oxygen, and strong ultraviolet radiation has brought challenges to the survival of animals and plants here. At the same time, the long-term adaptive evolution process has endowed these unique animals and plants with unique functional advantages to meet the needs of survival. Therefore, the protection and development of the dominant anti-environmental stress functional genes of animals unique to extreme environments is of extremely important value. [0003] The pika is an endemic and key species of the Qinghai-Tibet Plateau, which plays an important role in maintaining the balance and biodiversity of the Qing...
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