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Accurate and Phenol Free DNA Sexing of Day 30 Porcine Embryos by PCR
Published on: February 14, 2016
A rapid method for sexing the bovine embryo
Khairy M A Zoheir1, Ahmed A Allam
1National Research Centre, Cell Biology Department, Dokki, Cairo, Egypt. khma25@gmail.com
Animal Reproduction Science
|January 12, 2010
Summary
This study developed a rapid, accurate, and cost-effective method for bovine embryo sexing using loop-mediated isothermal amplification (LAMP). The technique allows naked-eye detection, simplifying sex determination in cattle breeding programs.
Area of Science:
- Animal Science
- Molecular Biology
- Reproductive Technology
Background:
- Accurate sex determination of bovine embryos is crucial for efficient cattle breeding and herd management.
- Traditional methods for embryo sexing can be time-consuming, expensive, and require specialized equipment.
- Loop-mediated isothermal amplification (LAMP) offers a potential alternative for rapid DNA amplification under constant temperature.
Purpose of the Study:
- To develop and validate an improved, field-applicable embryo sexing technique for cattle using LAMP.
- To assess the economic viability and ease of use of the new method compared to existing techniques.
- To enable precise manipulation of offspring sex ratios in bovine breeding programs.
Main Methods:
- DNA was extracted from bovine embryo biopsies (1-5 cells) for sex determination via LAMP.
- Ethidium bromide or CuSO(4) was added to LAMP products, inducing visible color change and precipitate for naked-eye detection.
- Sex results were confirmed using polymerase chain reaction (PCR) on remaining embryonic tissue.
- The entire procedure, including biopsy and sexing, was completed within approximately 45 minutes.
Main Results:
- The LAMP-based method achieved 100% accuracy in sex prediction for bovine embryos when using more than three dissociated blastomeres.
- The addition of ethidium bromide or CuSO(4) allowed for visual sex determination without electrophoresis or turbidity meters.
- The procedure was efficient, with a total time of about 45 minutes for sexing 58 bovine embryos.
- The method demonstrated high specificity and sensitivity for bovine DNA amplification.
Conclusions:
- The developed LAMP-based embryo sexing technique is a rapid, accurate, and cost-effective tool for bovine breeding.
- This method is suitable for field applications due to its simplicity and lack of requirement for specialized equipment.
- The technique facilitates the manipulation of sex ratios in cattle, offering significant advantages for breeding programs.
