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United States Department of Agriculture

Agricultural Research Service

Research Project: Genetic Characterization, Genetic Improvement, and Best Horticultural Management Practices for Subtropical/Tropical Ornamental Germplasm

Location: Subtropical Horticulture Research

Title: Conserved genetic regions across angiosperms help develop low copy nuclear markers in gymnosperms: an example using cycads

Authors
item Salas-Leiva, Dayana -
item Meerow, Alan
item Francisco-Ortega, Javier -
item Calonje, Michael -
item Griffith, Patrick -
item Stevenson, Dennis -
item Nakamura, Kyoko

Submitted to: Molecular Ecology Resources
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: January 20, 2014
Publication Date: N/A

Interpretive Summary: Several individuals of the Caribbean Zamia group (Zamia pumila complex) and other cycads were used to identify potential low copy nuclear genes for further studies of the evolutionary relationships within the complex and also among genera of the order Cycadales. Two strategies were used to select target loci: 1) a search of the highly conserved Arabidopsis sequence (COS) set and, 2) a search of the Arabidopsis-Populus-Oryza-Vitis Shared Single Copy genes (APVO SSC) genes. These two data sets were queried against the expressed sequence tag (EST) libraries for Zamia available in Genebank. From the first strategy, 30 loci were selected, and from the second, 16 loci. In both cases the matching Genbank accessions of Zamia sequences were used as a query for retrieving highly similar sequences from Cycas, Picea or Pinus species. A total of 29 loci were successfully amplified as a single band of which 20 were low copy loci. These loci showed different diversity and divergence levels. A preliminary screening allowed us to select 8 promising loci for the Z. pumila complex and 4 loci for all cycads.

Technical Abstract: Several individuals of the Caribbean Zamia clade (Zamia pumila complex) and other cycads were used to identify potential low copy nuclear genes for further studies of the phylogeographic relationships within the Caribbean Z. pumila complex and also among genera of Cycadales. Two strategies were used to select target loci: 1) a tblastX search of Arabidopsis conserved ortholog sequence (COS) set and, 2) a tblastX search of Arabidopsis-Populus-Oryza-Vitis Shared Single Copy genes (APVO SSC) genes against the EST Zamia databases in Genebank. From the first strategy, 30 loci were selected, and from the second, 16 loci. In both cases the matching Genbank accessions of Zamia were used as a query for retrieving highly similar sequences from Cycas, Picea or Pinus species. After retrieving and aligning all the sequences in each locus, intron predictions were completed and used to assist in primer design. PCR was carried out in three rounds to detect paralogous loci. A total of 29 loci were successfully amplified as a single band of which 20 were low copy loci. These loci showed different diversity and divergence levels. A preliminary screening allowed us to select 8 promising loci (40S, ATG2, BG, GroES, GTP, LiSH, PEX4 and TR) for the Z. pumila complex and 4 loci (COS26, GroES, GTP and HTS) for Cycadales.

Last Modified: 4/20/2014
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