Isozyme Variation within and among Accessions of Annual Phalaris Species in North American Germplasm Collections Maria Matus and Pierre Hucl* ABSTRACT Accessions of annual Phalaris species in germplasm collections lack adequategenetic characterization for utilization. The objective of this study wasto assess the level of isozyme variability of five annual Phalaris species, including 49 annual canarygrass (P. canariensis L.; 2n = 2x = 12), 24 P. brachystachys Link. (2n = 2x = 12), 48 P. minor Retz. (2n = 4x = 28), 19 P. paradoxa L. (2n = 2x = 14), and P. angusta Nees ex Tris (2n = 2x = 14) accessions fromNorth Ameri- can germplasm collections. Eight enzymeswere used to detect poly- morphism within and among accessions by scoring the presence or absence of 62 bands. Polymorphism was detected in 6 of 19 bands in P. canariensis, 12 of 22 bands in P. brachystachys, 34 of 40 bands in P. minor, 19 of 29 bands in P. paradoxa, and 6 of 21 bands in P. angusta. Polymorphism was detected in 23 accessions (47%) P. canariensis, 15 accessions (63%) ofP. brachystachys, 35 accessions (73%) of P. minor, four accessions (19%) of P. paradoxa and three accessions of P. angusta (100%). Within-accession polymorphic indi- ces ranged from0 to 0.03 in P. canariensis, 0 to 0.02 in P. brachysta- chys, 0 to 0.06 in P. minor, 0 to 0.04 in P. paradoxa, and 0.01 to 0.04 in P. angusta. Jaccard’ssimilarity coefficients ranged from0.83 to 1.0 in P. canariensis, 0.78 to 1.0 in P. brachystachys, 0.47 to 1.0 in P. minor, 0.44 to 1.0 in P. paradoxa, and 0.81 to 0.86 in P. angusta. Little isozyme variation existed either within or among accessions of the four diploid species. Phalaris minor, an autotetraploid, possessed greater variation among accessions rather than within accessions. In summary, limited variation was detected by means of isozymes in these five annual Phalaris species obtained from North American germplasm collections. ,ANNUAL CANARYORASS, known as canaryseed, is the only annual species of the genus Phalaris that has gained commercial importance (Anderson, 1961). Among the other annual species, P. angusta (Timothy canarygrass) is used as a fodder crop. Phalaris minor, P. brachystachys, and P. paradoxa occur as weeds in cultivated crops. These five annual species are predomi- nantly self-pollinated (Carlson et al., 1996). Argentina, Morocco, and Australia have been the traditional world producers of canarygrass as a source of birdseed. Over the last decade, Canadian production of annual ca- narygrass seed has averaged 112000 metric tons per year, approximately 65 % of world seed production (Put- manet al., 1996). Currently, ’Keet’, ’Elias’, and ’CDC Maria’ are the only registered annual canarygrass culti- vars grown in Canada. Future crop improvements to annual canarygrass cultivars by either intraspecific or interspecific hybridizations, or mutagenesis to introduce new traits, may benefit from a better understanding of the genetic variability contained in annual Phalaris germplasm collections. In the absence of intraspecific variation, strategies for annual canarygrass improve- ment mayinclude interspecific hybridization; however, this technique has limited potential in the short term because of crossing barriers. Phalaris brachystachys should be the best candidate for hybridization with P. canariensis because both share the same chromosome number (Anderson, 1961). The genetic relationship among the annual species P. canariens&, P. brachystachys, P. minor, P. paradoxa, and P. angusta is thought to be divergent on the basis of speculation that each species evolved along different lines of evolution (Anderson, 1961). Four variations sterile floret structure have evolved in Phalaris and have been used to bolster speculation on the relationships amongthe 10 annual and five perennial species in the genus. Lines of development from a hypothetical primi- tive Phalaris, include (i) P. brachystachys and P. truncata with reduced and swollen sterile florets, (ii) P. cana- riensis with bract-like and chaffy sterile florets, (iii) P. paradoxa and P. coerulescens with obsolete sterile florets, and (iv) the other 10 species in the genus with subulate sterile florets. The species with subulate sterile florets are further divided into three lines of develop- ment on the basis of geographic distribution: Old World species (P. minor, P. aquatica, and P. maderensis ), cir- cumboreal species (P. arundinacea), and NewWorld species (P. californica, P. caroliniana, P. amethystina, P. lemmonii, P. platensis, and P. angusta). Further divi- sions can be made among the New World species on the basis of their cytology and inflorescence and spike- let morphology. Isozymes have been used extensively to characterize accessions in various crop germplasm collections (Ynd- gaard and Hoskuldsson, 1985), but their application in Phalaris germplasm collections is limited (Villarroya et al., 1983). The objective of this study was to assess the level of variability within and amongNorth American accessions of P. canariensb, P. brachystachys, P. minor, P. paradoxa, and P. angusta by means of eight isozymes. MATERIALS AND METHODS Phalaris Germplasm and Sample Preparation The geographic origin of 143accessionsof Phalaris, includ- ing 49 P. canariensis, 48 P. minor, 24 P. brachystachys, 19 M. Matus, Dep. of Plant Sciences, Univ. of Saskatchewan, Saskatoon, 51 Campus Dr., SK, S7N 5A8, Canada; and P. Hucl, Crop Develop- ment Center, Univ. of Saskatchewan, 51 CampusDr., Saskatoon, SK, S7N 5A8, Canada. Received 29 April 1998. *Corresponding author (hucl@sask.usask.ca). Published in Crop Sci. 39:1222-1228 (1999). Abbreviations: ACO,aconitase; AAT, aspartate aminotransferase; CDC, Crop Development Centre; LAP, leucine aminopeptidase; MDH, malate dehydrogenase; p~, frequency of the ith band; PGD, phosphogluconate dehydrogenase; PGI, phosphoglucoisomerase; PX, polymorphic index for a given enzyme and accession; PXw, within- accession polymorphic index; SKD, shikimate dehydrogenase; TPI, triose phosphate isomerase. 1222 Published July, 1999