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Methods for Injecting Avian Eggs

a technology of avian eggs and injection methods, which is applied in the field of methods for injecting avian eggs, to achieve the effects of reducing morbidity and mortality, reducing embryo trauma, and improving viability and hatching rates

Inactive Publication Date: 2008-01-31
EMBREX INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0007] The present invention provides a method of manipulating avian eggs containing embryos, in particular early stage embryos, which may result in reduced trauma to the embryo and improved viability and hatch rates. Accordingly, the present invention provides improved methods of inserting or implanting a variety of devices (e.g., a delivery device, sampling device, and / or a detector device, and the like) into an avian egg containing an embryo, in particular an early embryo (e.g., a blastoderm), which may result in lower levels of morbidity and mortality among the manipulated embryos. The methods of the invention are particularly useful in methods of delivering vaccines, vitamins, growth promoting hormones and growth factors, enzymes, cytokines, nucleic acids, and / or cells to a bird in ovo. For example, the method may be used to produce a chimeric bird (i.e., containing foreign cells) or a transgenic bird (i.e., containing a foreign nucleic acid sequence). The methods of the invention are also useful for collecting samples or information from a bird in ovo, for example, for use in methods of gender sorting, determining embryo viability, and / or obtaining information about the genetic profile of the embryo.

Problems solved by technology

Current methods of manipulating bird eggs containing early embryos may be undesirable because they may result in unacceptably low hatch rates.

Method used

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  • Methods for Injecting Avian Eggs
  • Methods for Injecting Avian Eggs
  • Methods for Injecting Avian Eggs

Examples

Experimental program
Comparison scheme
Effect test

example 1

[0117] The following is an exemplary method used to inject fluid or cells into recipient chicken blastoderms in ovo through the blunt end of the egg: [0118] 1. Surface sterilize the entire egg surface of Day E 0 eggs by wiping with 100% ethanol. [0119] 2. Candle the egg to locate the air cell. The position of the air cell is marked on the shell with a pencil. [0120] 3. A window (small opening) is made at the blunt end of the egg (in center of air cell) by dremeling away the shell. Care is taken to keep the outer shell membrane intact.

All of the following steps are preferably carried out in a laminar hood or in a clean room: [0121] 4. The blunt end of the egg, where the outer shell membrane is exposed through the window in the shell, and the surrounding area are again surface sterilized using 70% ethanol. [0122] 5. The outer shell membrane is carefully removed with forceps and scalpel (#11 blade; Personna Medical) taking care not to leave any jagged edges to the membrane. [0123] 6....

example 2

[0128] A series of experiments was carried out to evaluate the use of manual injection as described in Example 1 to inject either fluid or cells into the subgerminal cavity of recipient chicken blastoderms (approximately Stage X) in ovo. Donor cells were also from Stage X chicken blastoderms. In some cases, the recipient embryo was compromised prior to injection. In early experiments, a small droplet (a few μl) of an albumen solution was deposited on the inner shell membrane above the blastoderm and injection was performed by piercing the membrane through the albumen droplet. In later experiments, this step was omitted. Eggs were either not stored or stored from 4 to 8 days prior to treatment.

[0129] Hatch rates in control and manipulated eggs were determined. The rate of successful chimera production was also evaluated. The results are shown below in Table 1.

TABLE 1Percent Chimera ObtainedRecipientCompromiseDonorNo. ofNo. ofHatchability ofHatchabilityBased onBased onEggRecipientE...

example 3

[0130] Certain methods of physical compromise of the embryo, such as ultraviolet (UV)-irradiation or laser ablation, may be administered through a window in the egg shell and shell membranes. We have developed a method to create a window in the side of the egg, seal it, and incubate the egg under standard conditions (99 F, rocking), that allows a very high percentage (>90%) of embryos to develop normally at two days (FIG. 7). This method may be used to evaluate the effects of compromise and chimera production at two days of embryonic development. Further, this method may be used to inject into the side of the egg. Briefly, the eggs were treated as follows: [0131] 1. Eggs were stored at 60 F, 75% humidity for 0-7 days, on their long axis, to steer the blastoderm towards the center of the side of the egg. [0132] 2. The position facing upwards was marked with a sharpie pen. [0133] 3. Following storage, a 7-mm hole was drilled at the side of the egg, at the location of the pen mark. [01...

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Abstract

The present invention provides improved methods of injecting an avian egg containing an embryo, preferably an early embryo (e.g., a blastoderm). The methods of the invention may be used to deliver a substance to an egg, remove a sample from an egg, and / or to insert a detector device into an egg to collect information therefrom. In preferred embodiments, the invention is used to deliver a substance to the embryo in ovo. In other preferred embodiments, the invention is used to produce chimeric or transgenic avian embryos in ovo.

Description

RELATED APPLICATION INFORMATION [0001] This application is a divisional of U.S. application Ser. No. 10 / 216,427, filed Aug. 9, 2002 (allowed), which claims the benefit of U.S. Provisional Application No. 60 / 312,015 filed Aug. 13, 2001, which is incorporated by reference herein in its entirety.FILED OF THE INVENTION [0002] The present invention relates to methods of manipulating an egg containing an embryo and, in particular, to methods of introducing material into or removing material from an egg containing an avian embryo. BACKGROUND OF THE INVENTION [0003] There are a number of applications for which it is desirable to inject eggs containing early avian embryos. For example, it may be desirable to deliver a substance to an early embryo, such as a blastoderm. To illustrate, it may be desirable in the poultry industry to manipulate an early embryo in ovo to introduce a foreign nucleic acid molecule (i.e., to create a transgenic bird) or to introduce a foreign cell(s) (i.e., to creat...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): A01K45/00A01K67/027C12N15/09C12N15/85
CPCA01K45/007A01K67/0271C12N15/8509A01K2227/30A01K2267/03A01K2217/05
Inventor MENDU, NANDINIBLAND, MOLLYWOLFE, STEPHENHEBRANK, JOHNMAHATO, DIPAK
Owner EMBREX INC
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