Wolf Prize in Agriculture (1157 Pages)
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- Oat–maize chromosome addition lines: A new system for mapping the maize genome (corn
- ABSTRACT Novel plants with individual maize chromo- somes added to a complete oat genome have been recovered via embryo rescue from oat (
960 Wolf Prize in Agriculture 209. Rines, H.W., R.L. Phillips, R.G. Kynast, R.J. Okagaki, W.E. Odland, A.O. Stec, M.S. Jacobs, and S.R. Granath. 2005. Maize chromosome additions and radiation hybrids in oat and their use in dissecting the maize genome. In: In the wake of the double helix: from the green revolution to the gene revolution. Eds. Tuberosa, R., R.L. Phillips, and M. Gale, Avenue Media, Bologna, Italy, pp. 427-442. 210. Tuberosa, R., R.L. Phillips, and M. Gale. (Eds.) 2005. In the wake of the double helix: From the green revolution to the gene revolution. Avenue Media, Bologna, Italy. 772 pp. CD produced in 2006. 211. Rines, H.W., S.J. Molnar, N.A. Tinker, and R.L. Phillips. 2006. Oat. In: Kole, C. (ed.). Genome Mapping and Molecular Breeding in Plants: Cereals and Millets Vol. 1. Springer, Inc., NY, USA. pp. 211-242. 212. Phillips, R.L. 2006. Genetic tools from nature and the nature of genetic tools. In: CSSA Golden Anniversary Symposium. Ed. C. Stuber. Crop Sci. 46:2245-2252. 213. Phillips, R.L., W.E. Odland, and A.L. Kahler. 2007. Rice as a reference genome and more. In: 5 th Intl. Rice Genetics Symp., Eds. D.S. Brar, D. Mackill, and B. Hardy. pp. 3-15. 214. Phillips, R.L., and H.W. Rines. 2008. Genetic analyses with oat maize addition and radiation hybrid lines. In: The Maize Handbook: Domestication, Genetics, and Genome, Eds. J.L. Bennetzen and S.C. Hake, Springer-Life Sciences, New York, NY (in press). 38_2006-7 Phillips.p65 06-Mar-09, 7:49 PM 960
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Proc. Natl. Acad. Sci. USA Vol. 94, pp. 3524–3529, April 1997 Agricultural Sciences
͞chromosome-specific͞cosmid library͞repetitive DNA͞cloning) E VGUENI
V. A NANIEV
*, O SCAR
R IERA
-L IZARAZU
*, H OWARD
W. R INES
* † , AND R ONALD L. P HILLIPS
* ‡ *Department of Agronomy and Plant Genetics and Plant Molecular Genetics Institute, University of Minnesota, 1991 Upper Buford Circle, St. Paul, MN 55108; and † Plant Sciences Research Unit, U.S. Department of Agriculture–Agricultural Research Service, St. Paul, MN 55108 Contributed by Ronald L. Phillips, February 10, 1997 ABSTRACT Novel plants with individual maize chromo- somes added to a complete oat genome have been recovered via embryo rescue from oat ( Avena sativa L., 2n ؍ 6x ؍ 42) ؋ maize ( Zea mays L., 2n ؍ 20) crosses. An oat–maize disomic addition line possessing 21 pairs of oat chromosomes and one maize chromosome 9 pair was used to construct a cosmid library. A multiprobe (mixture of labeled fragments used as a probe) of highly repetitive maize-specific sequences was used to selectively isolate cosmid clones containing maize genomic DNA. Hybridization of individual maize cosmid clones or their subcloned fragments to maize and oat genomic DNA revealed that most high, middle, or low copy number DNA sequences are maize-specific. Such DNA markers allow the identification of maize genomic DNA in an oat genomic background. Chi- meric cosmid clones were not found; apparently, significant exchanges of genetic material had not occurred between the maize-addition chromosome and the oat genome in these novel plants or in the cloning process. About 95% of clones selected at random from a maize genomic cosmid library could be detected by the multiprobe. The ability to selectively detect maize sequences in an oat background enables us to consider oat as a host for the cloning of specific maize chromosomes or maize chromosome segments. Introgressing maize chromo- some segments into the oat genome via irradiation should allow the construction of a library of overlapping fragments for each maize chromosome to be used for developing a physical map of the maize genome. Chromosome addition lines of different plant species (1–5) have been generated to introgress valuable genes from wild or cultivated relatives into host plant species. Alien chromosome additions have been used for gene mapping (2, 6, 7) and serve as an enriched source of markers for positional cloning and constructing physical maps of specific chromosomes. The discovery of maize-chromosome retention in oat ‘‘haploids’’ after oat ϫ maize crosses and the recovery of stable maize chromosome-addition oat lines (8, 9) should allow the devel- opment of a system of chromosome analysis similar to that available in mammalian hybrid-cell systems (10–12). Such a system may be used for gene assignment, isolation of chromo- some-specific probes (13), flow sorting (14) and microdissec- tion of chromosomes (15), development of chromosome- specific ‘‘paints’’ of fluorochrome-labeled DNA fragments (16–18), physical mapping, and selective isolation and map- ping of cDNAs of a particular chromosome (19, 20). In this paper we describe an approach for isolating clones containing large-fragment maize DNA of a single-chromo- some origin from the oat–maize chromosome addition lines that will assist in the construction of physical maps for maize chromosomes. The approach is based on cloning genomic DNA of an oat–maize chromosome-addition line in an appro- priate vector and subsequent use of maize-specific dispersed repetitive DNA sequences as detection probes to isolate clones carrying maize genomic DNA. Such an approach was success- fully applied to the isolation of human-specific DNA fragments in cosmid libraries constructed from DNA of human-rodent hybrid cell lines carrying individual human chromosomes. In those experiments the probes included Alu-repeats (21), a Cot1 DNA fraction (22), or labeled total human DNA (23). Plant repetitive sequences that are apparently species- specific were first isolated from rye (24, 25) and later found in many other species (26). Comparative studies revealed a strong correlation between the proportion of species-specific re- peated families in a genome and phylogenetic relationships (27, 28). About 90–95% of all randomly tested genomic repeated sequences from barley were detected in wheat and rye (27). According to DNA reassociation studies, less-related species such as maize and wheat have less than 10% nucleotide sequences in common (28). The success of isolating maize DNA from oat–maize chromosome-addition lines depends on how many maize-specific (relative to oat) high-copy-number dispersed nucleotide sequences are found in the maize ge- nome. Genome analysis in grasses, including maize and oat, reveals that a major portion of genomic DNA consists of families of repetitive sequences dispersed throughout the genome (26, 29, 30). According to DNA–DNA renaturation data in maize (28), unique sequences of average length 2,100 bp are interspersed with mid-repetitive sequences. Direct analysis of sequences adjacent to several maize genes reveals that these genes are flanked by highly repetitive DNA se- quences (31). Thus, maize large DNA inserts cloned in an appropriate vector will likely carry some kind of dispersed maize-specific nucleotide sequence. A cosmid vector was chosen for this project because of the high cloning efficiency and relatively large insertion size. Because the oat genome is about 11,300 Mb, an added maize chromosome with median size of about 250 Mb would constitute about 2–4% of the total nuclear DNA of an oat–maize chromosome addition line. We found that the major part of the maize genome consists of nucleotide sequences that do not cross-hybridize to oat genomic sequences under standard hybridization conditions. This predominant nonhomology between oat and maize genomic sequences means that many maize genomic sequences can be directly used for Southern blot hybridization on DNA from oat–maize chromosome addition lines to selectively detect sequences of maize origin. A mixture of highly repetitive dispersed DNA sequences of maize was used as a maize- specific multiprobe to screen a cosmid library of the chromo- some addition line. A group of maize-specific cosmids with The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked ‘‘advertisement’’ in accordance with 18 U.S.C. §1734 solely to indicate this fact. Copyright ᭧ 1997 by T HE N
A CADEMY OF S CIENCES OF THE USA 0027-8424 ͞97͞943524-6$2.00͞0 PNAS is available online at http: ͞͞www.pnas.org. Abbreviation: RFLP, restriction fragment length polymorphism; Mb, megabase(s). ‡ To whom reprint requests should be addressed. 3524 Copyright (1997) National Academy of Sciences, USA. 38_2006-7 Phillips.p65 06-Mar-09, 7:49 PM 975
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