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Method for improving bovine in-vitro fertilization efficiency

A bovine body and in vitro technology, applied in bovine in vitro fertilization, the application of related working test solution in bovine in vitro fertilization, in the field of agriculture, animal husbandry and veterinary reproduction, can solve the problem of limited efficiency of in vitro fertilization and so on

Active Publication Date: 2021-03-16
天津力牧生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0015] However, the efficiency of bovine frozen semen for in vitro fertilization with oocytes remains limited

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0064] Embodiment 1: the culture method of bovine in vitro fertilization embryo (ex vivo collection)

[0065] In the present embodiment, in step (1), the collected oocytes were placed in transport medium containing HEPES at 38.8° C. without carbon dioxide supply, and transported back to the laboratory within 24 hours.

[0066] 1. Reagents

[0067] In the specific test of the present invention, if not otherwise specified, the relevant reagents used are described in detail as follows:

[0068] The transportation culture fluid containing HEPES used in this example contains: glycine 50.0mg / L, L-alanine 25.0mg / L, L-arginine hydrochloride 70.0mg / L, L-aspartic acid 30.0mg / L, L-cystine dihydrochloride 26.0mg / L, L-glutamic acid 75.0mg / L, L-glutamine 100.0mg / L, L-histidine hydrochloride monohydrate 21.88mg / L, L-hydroxyproline 10.0mg / L, L-isoleucine 40.0mg / L, L-leucine 60.0mg / L, L-lysine hydrochloride 70.0mg / L, L-methionine 15.0mg / L, L-phenylalanine 25.0mg / L, L-proline 40.0mg / L, L-se...

Embodiment 1A

[0106] Embodiment 1A: the culture method of bovine in vitro fertilization embryo (ex vivo collection)

[0107] With reference to the method of Example 1, tests were carried out on three kinds of cattle, Holstein (dairy breed), Simmental (beef breed), and Chinese buffalo (serving breed), respectively. The results showed that the cleavage rate was 85-90%. Within the range, the morula rate was in the range of 62-67%, the blastocyst rate was in the range of 50-55%, the apoptosis rate was in the range of 4-6%, and the blastocyst hatching rate on the 9th day was 72-5%. Within the range of 77%; the maturation rates of oocytes in vitro maturation of Holstein, Simmental, and Chinese buffalo were 83.3%, 80.6%, and 75.6%, respectively.

Embodiment 1B

[0108] Embodiment 1B: the culture method of bovine in vitro fertilization embryo (in vitro collection)

[0109] With reference to the method of Example 1, four kinds of cattle were tested respectively for Chinese yellow cattle (Nanyang cattle, service breed), Holstein cattle (dairy breed), Simmental cattle (beef breed), and Chinese water buffalo (service breed), However, the formulations of the fertilization culture medium and the semen preparation culture medium were respectively modified, and the remaining test fluids and operations remained unchanged. The formula of the fertilization medium was changed to: containing 112.0mM sodium chloride, 4.02mM potassium chloride, 2.25mM calcium chloride dihydrate, 0.52mM magnesium chloride hexahydrate, 0.83mM potassium dihydrogen phosphate, 37.0mM sodium bicarbonate, 1.25 mM sodium pyruvate, 10μg / ml heparin, 4mg / ml bovine serum albumin (BSA), 100U / ml penicillin, 100μg / ml streptomycin, 10umoL / L α-lipoic acid, 25μg / ml folic acid, 2.5mg...

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Abstract

The invention relates to a method for improving the bovine in-vitro fertilization efficiency. On one hand, the invention relates to a bovine in-vitro fertilization embryo culture method. The method comprises the following steps of performing collection and in-vitro maturation of oocytes, performing in-vitro fertilization by using a fertilization culture solution and a seminal fluid preparation culture solution, and performing in-vitro embryo culture and preservation. A maturation culture solution used in the method is a BY basic culture solution added with 100 mL / L of FBS, 10 [mu]g / mL of FSH,10 [mu]g / mL of LH, 1 [mu]g / mL of E2 and 20 ng / mL of EGF, and the used fertilization culture solution and seminal fluid preparation culture solution contain sodium pyruvate, heparin and bovine serum albumin. The invention also relates to a bovine in-vitro fertilization method, and also relates to the fertilization culture solution and the seminal fluid preparation culture solution used in the methods. The method using a reagent shows the excellent technical effect of significantly improving the in-vitro fertilization efficiency.

Description

technical field [0001] The invention belongs to the technical field of animal reproduction, and relates to a technology for agricultural-animal husbandry and veterinary reproduction, in particular to a method for bovine in vitro fertilization. In addition, the present invention also relates to the use of related working test solutions used in bovine in vitro fertilization. Application in fertilization. The bovine in vitro fertilization method of the invention has excellent technical effects. Background technique [0002] In Vitro Fertilization (In Vitro Fertilization) or (external fertilization) refers to the technology in which mammalian sperm and eggs complete the fertilization process in an artificially controlled environment outside the body. It is referred to as IVF in English. Because it is inseparable from embryo transfer technology (ET), it is also referred to as IVF-ET. In biology, animals obtained by transplanting in vitro fertilized embryos into mothers are call...

Claims

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Application Information

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IPC IPC(8): C12N5/075C12N5/073
CPCC12N5/0609C12N5/0604C12N2501/31C12N2501/11C12N2501/392C12N2500/32C12N2500/33C12N2500/38C12N2500/35C12N2500/14C12N2500/24C12N2500/16C12N2500/12C12N2500/40C12N2500/36C12N2500/34C12N2501/999C12N2501/105C12N2500/30C12N2501/91
Inventor 王娜王小武郝少强赵明礼韩勇权郭春明许晓椿
Owner 天津力牧生物科技有限公司
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