Analysis of the CHD gene in birds as a molecular marker for sex determination

Abstract

Sex determination in birds is extremely difficult, primarily due to the fact that more than 50% of bird species are monomorphic. Many traditional methods that have been used before defining molecular genetics methods have their biggest lack as their main characteristic – the unreliability. The results obtained using different methods should be primarily reliable and analysis should be fast, secure and cost-effective. Highly conserved CHD gene has been discovered in 1995 on the W chromosome, and in 1997 on the Z chromosome. The difference in the intron length between the two chromosomes allows sex determination after the PCR reaction. Blood, feathers, buccal swabs, faeces and formalin fixed paraffin embedded tissue samples were used for analysis. We compared the success of the reaction depending on which sample was used, also we compared the success of the reactions when used ordinary polymerases and polymerase designed to be resistant to PCR inhibitors. Using PCR and set of primers 2550F/2718R we have examined the possibility of using CHD gene as molecular markers for sex determination of 550 individuals originating from 83 species of birds widely distributed along the phylogenetic tree. Sex was successfully determined in 74 bird species. In 26 of those it was carried out for the first time using molecular markers and PCR...

Određivanje pola kod ptica je izuzetno teško, prvenstveno zbog činjenice da je preko 50% vrsta monomorfno. Pre definisanja molekularno genetičkih metoda, za determinaciju pola kod ptica koristile su se mnoge tradicionalne metode čija je glavna karakteristika, a ujedno i najveća mana – nepouzdanost. Rezultati koji se dobijaju primenom različitih metoda moraju biti pre svega pouzdani, a analize ekonomične, brze i bezbedne. Visokokonzervisani CHD gen je otkriven 1995. godine na W hromozomu ptica, dok je na Z hromozomu otkriven 1997. godine. Postojanje razlike u dužini introna između Z i W hromozoma omogućava razlikovanje polova nakon PCR reakcije. U radu su kao uzorci korišćeni perje, krv, bris usne duplje, feces i parafinski tkivni isečci različitih tkiva. Poređena je uspešnost reakcija u zavisnosti od toga koji je uzorak korišćen, kao i uspešnost reakcija kada se koristila obična polimeraza i polimeraza specijalno dizajnirana da bude otporna na inhibitore PCR reakcije. Korišćenjem PCR tehnike i seta prajmera 2550F/2718R ispitana je mogućnost primene CHD gena kao molekularnog markera za određivanje pola kod 550 jedinki poreklom od 83 vrsta ptica široko distribuiranih duž filogenetskog stabla...