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Detection of single copy sequences using BAC-FISH and C-PRINS techniques in sunflower chromosomes

PAOLA TALIA1 , EDUARDO J. GREIZERSTEIN1 , H. ESTEBAN HOPP1,2 , NORMA PANIEGO1 , LIDIA POGGIO2 AND RUTH A. HEINZ1,2*

1. Instituto de Biotecnología, CICVyA, INTA Castelar, N. Repetto y Los Reseros s/n, 1686 Hurlingham, Provincia de Buenos Aires, Argentina.
2. Departamento de Ecología, Genética y Evolución and Departamento de Fisiología, Biología Molecular y Celular, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad Universitaria, Pabellón II, C1428EHA, Buenos Aires, Argentina.

*Address correspondence to: Ruth Heinz. Instituto de Biotecnología CICVyA / INTA. Dr. N. Repetto y Los Reseros. (1686) Hurlingham, Buenos Aires. Argentina. Fax: (54-11) 4481 2975. Tel.: (54-11) 4621 1447 ó 0199. E-mail: email

BIOCELL 2011, 35(1), 19-28. https://doi.org/10.32604/biocell.2011.35.019

Abstract

Bacterial artificial chromosome - fluorescence in situ hybridization (BAC-FISH) and cyclingprimed in situ labeling (C-PRINS) techniques were evaluated for integration of physical and genetic maps of sunflower (Helianthus annuus L.). Single-site SSR markers were selected from three linkage groups of a high-density sunflower genetic map. This selection was based on previously identified QTL associated to S. sclerotiorum. These markers were used to select BACs contaning single copy sequences for BAC-FISH aplication. Blocking of highly dispersed repetitive sunflower sequences reduced unspecific hybridization, and allowed the detection of specific signals for BACs containing SSR markers HA4222 and HA2600, anchored to LG 16 and LG 10, respectively. Single-site FISH signal detection was optimized by adjusting the relative quantity and quality of unlabelled repetitive sequences present in the blocking DNA. The SSR marker ORS1247 anchored to the LG 17 was detected by C-PRINS, which yielded fluorescence signals that were specific and intense. This progress in localizing single-copy sequences using BAC-FISH and indirect CPRINS strategies in sunflower will facilitate the integration of genetic and physical maps, allowing the identification of

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TALIA, P. (2011). Detection of single copy sequences using BAC-FISH and C-PRINS techniques in sunflower chromosomes. BIOCELL, 35(1), 19–28.

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