Free Access
Volume 37, Number 6, November-December 2006
Page(s) 687 - 698
Published online 20 October 2006
Apidologie 37 (2006) 687-698
DOI: 10.1051/apido:2006043

An optimized method for the generation of AFLP markers in a stingless bee (Melipona quadrifasciata) reveals a high degree of intracolonial genetic polymorphism

Gustavo R. Makerta, Robert J. Paxtonb and Klaus Hartfelderc

a  Depto. Genética, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, Brazil
b  School of Biological Sciences, Queen's University Belfast, Belfast, UK
c  Depto. Biologia Celular e Molecular e Bioagents Patogênicos, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, Brazil

(Received 22 June 2005 - Revised 26 February 2006 - Accepted 28 February 2006 - published online 20 October 2006)

Abstract - This study describes an optimized protocol for the generation of Amplified Fragment Length Polymorphism (AFLP) markers in a stingless bee. Essential modifications to standard protocols are a restriction enzyme digestion (EcoRI and Tru1I) in a two-step procedure, combined with a touchdown program in the selective PCR amplification step and product labelling by incorporation of $\alpha $[ 33P] dATP. In an analysis of 75 workers collected from three colonies of Melipona quadrifasciata we obtained 719 markers. Analysis of genetic variability revealed that on average 32% of the markers were polymorphic within a colony. Compared to the overall percentage of polymorphism (44% of the markers detected in our bee samples), the observed rates of within-colony polymorphism are remarkably high, considering that the workers of each colony were all offspring of a singly mated queen.

Key words: amplified fragment length polymorphism / genotyping / stingless bee / genetic polymorphism / Melipona / Apidae

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© INRA, DIB-AGIB, EDP Sciences 2006