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United States Department of Agriculture

Agricultural Research Service

Title: Construction of a Sugarbeet Bac Library from a Hybrid That Combines Diverse Traits

Authors
item McGrath, J Mitchell
item Shaw, R Scott
item DE Los Reyes, Benildo - UNIVERSITY OF MAINE
item Weiland, John

Submitted to: Plant Molecular Biology Reporter
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: March 15, 2003
Publication Date: April 1, 2004
Citation: McGrath, J.M., Shaw, R.S., De Los Reyes, B., Weiland, J.J. 2004. Construction of a sugarbeet BAC library from a hybrid that combines diverse traits. Plant Molecular Biology Reporter. 22(1):23-28.

Interpretive Summary: The sugar beet genome contains approximately 750 million nucleotide base pairs of DNA. Encoded in this DNA are sequences that contribute to the agronomic performance and economic return of the crop to growers. Identifying these sequences is a major challenge. A Bacterial Artificial Chromosome library was constructed containing 38,000 pieces of sugarbeet DNA, each piece about 100,000 nucleotide base pairs long, yielding a total number of pieces in excess of five times the DNA in a single sugar beet cell. This redundancy is necessary to increase the probability of recovering any randomly isolated clone (piece) in the library. The probability of recovering any clone of interest was assessed using known genes. Results showed the library contained all genes tested, and therefore the library is representative of the sugar beet genome. This library is an important resource that will allow sugar beet geneticists access to the specific genes and their sequences that are involved in agronomic performance. Knowing these sequences will allow breeders to select for genetic variation associated with increased agronomic performance.

Technical Abstract: A Bacterial Artificial Chromosome library with five-fold coverage of the ca. 750 Mb sugar beet genome was constructed from HinDIII-digested DNA from hybrid USH20, with an average insert size of 100 to 125 kb. Filter arrays were prepared that contained all clones and were used to assess the abundance and distribution of particular types of nucleotide sequences via filter-hybridization approaches. Using a ribosomal RNA gene probe, 1.2% of the library carried sequences similar to these highly repetitive, highly conserved sequences. A simple sequence repeat element (CA)8 thought to be predominantly distributed throughout centromere regions of all chromosomes was present in 1.6% of clones. Extra-nuclear DNA contamination was low, but chloroplast DNA contamination was greater than mitochondrial DNA (1.6 % of clones vs. 0.01%, respectively). Twenty-eight randomly chosen ESTs were screened against nylon filter arrays of the BAC library. For over half of the ESTs used as probes, a greater than expected number of hybridization signals were observed for a single copy sequence assuming 5X genome coverage, suggesting that many genes are duplicated in the beet genome.

Last Modified: 11/22/2014
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