The paper “ Bird's Sex Identification Using DNA Markers” is a meaty example of a lab report on biology. DNA was isolated from three types of samples these are blood, feathers, and muscle tissue. The collection of blood as a sample for DNA extraction could be from a live bird or a dead one. The sample should be fresh to maintain high quality and quantity of DNA and to prevent contamination. Fresh blood samples can be stored in 95% ethanol at room temperature or frozen. Feather samples could be obtained by plucking the bird and these provide a less painful collection method than drawing blood.
The feathers should be stored in a sterile sample bag immediately they are plucked from the bird to avoid contamination. Muscle tissues are extracted from dead birds and immediately frozen for the preservation of the quality. When freezing of tissues is not possible, one can place minced tissues in a vial of buffer (Seutin at el. 1991) or place small pieces into 95% ethanol for dehydration. Ultra-cold freezing represents the closest thing to a museum-quality tissue preservation method. Generally, determination of the sex of birds is quite difficult before puberty but in monomorphic species, it is difficult to even after puberty.
Chickens are difficult to sex morphologically hence the use of PCR for sex identification. The Z and W sex chromosomes evolved differently in birds from the mammalian X and Y chromosomes. In Chickens females are heterogametic and carry a copy of Z and W sex chromosomes but males are homogametic and carry 2 copies of the Z sex chromosome. The most critical question in sex identification of chickens is how the 2 types of sex chromosomes play a role.
It is not clear yet if female characteristics are developed by female-specific W chromosomes or male characteristics are defined by the dose of chromosome Z. Even though it is female-specific, the W chromosome has a similar structure to the mammalian male Y chromosome, except for its poorer gene structure, smaller size, and richness in heterochromatin and repeat sequences. There are 2 genes defined on the W chromosome. These are chromo helicase DNA binding protein (CHD1W) and ATP synthesis α -subunit (ATP5A1W).
Both genes are located in the non-recombined part of the W chromosome and their similar homologs (CHD1Z and ATP5A1Z) exist in the Z chromosome. This study aims to help breeders identify the sex of chickens without having to wait for the physical characteristics to manifest as they grow. The breeders normally have large numbers of birds and it would be tedious to physically inspect each bird for its sex. Method: Purification of genomic DNA from blood, muscle tissue, and feather sample. Protocol 1aThe purification of genomic DNA from preserved blood was as follows: after pipetting 20µ l of proteinase K into a sterile 1.5 ml microcentrifuge tube, 166µ l of PBS and 4µ l of RNase A was added into the tube.
A sample of preserved chicken blood was transferred using sterile forceps. This was incubated for 30 minutes at room temperature before adding 200µ l of buffer AL and then it was mixed thoroughly using vortexing for 15 seconds. A lid lock was placed on the tube and it was further incubated for 25-45 minutes. 200µ l of 95% ethanol (AR grade) was added to the sample and thoroughly mixed by vortexing for 15 seconds.
The DNA precipitated and was ready to be bound to the membrane.
List of references
Baquero A, Puerta A, Gutierrez G (2009). Magnitude Effects of Sexual Reinforcement in
Japanese quail (Coturnix japonica). Int. J. Comp. Psychol. 22: 113-126
Cerit H, Avanus K (2007). Sex determination by CHDW and CHDZ genes of avian sex
chromosomes in Nymphicus hollandicus. Turk. J. Vet. Anim. Sci. 31(6): 371-374.
Fridolfsson, A., and Ellegren, H. (1999) A simple and universal method for molecular sexing of
non-ratite birds. Journal of Avian Biology. 30, 116 – 121.
Freeland, J (2005) Molecular Ecology. Wiley. Chichester
Griffiths R. 2002. Sex identification using DNA markers. In: Molecular methods in
Ecology (BACKER A. J., Ed.), pp. 295–321, Blackwell Science, London.
Horvath, M. Martinez-Cruz, B. Negro, J. Kalmar, L and Goday, J. (2005). An overlooked
DNA source for non-invasive genetic analysis in birds. Journal of Avian Biology. 36, 84-88.