Korban Schuyler S, Wannarat Wannasiri, Rayburn Charlotte M, Tatum Tatiana C, Rayburn A Lane
Department of Natural Resources and Environmental Sciences, 310 ERML, 1201 W. Gregory Drive, University of Illinois, Urbana, IL 61801, USA.
Genome. 2009 Feb;52(2):148-55. doi: 10.1139/g08-109.
The genus Malus has anywhere between 25 and 33 species along with several subspecies. Malus species as well as clones within the same species have varying ploidy levels, as these are more than likely collected from different trees and (or) from different locations. In recent years, large numbers of Malus germplasm accessions have been collected and maintained at the United States National Germplasm Clonal Repository; however, genome sizes of this material have not yet been determined. In this study, leaf tissues from young grafted trees of 100 Malus species and hybrids growing in a nursery at the University of Illinois were collected and immediately used for extracting nuclei. Leaf tissues from apple and maize line W-22, used as an internal standard, were co-chopped and prepared for flow cytometric analysis. Apple nuclei were stained with propidium iodide, an intercalating dye, and a minimum of 8000 nuclei per sample were analyzed. Mean fluorescence of apple nuclei was then determined. A total of four replications per sample was used. Among 100 Malus accessions analyzed, one tetraploid, three triploid, and 96 diploid genotypes were identified. Significant differences in genome size were identified among the three ploidy types observed and also within diploid genotypes. The 2C mean value for tetraploids was 3.13 pg and ranged from 2.27 to 2.41 pg for triploids, whereas 2C values for diploids ranged between 1.44 and 1.72 pg. In addition, leaf impressions of young, fully expanded leaves were collected from young trees of 10 selected genotypes based on their ploidy and flow cytometric analysis and used to measure the nucleotypic parameter stomatal length. Ten stomata were measured per slide, three slides were analyzed per leaf, and three leaves were analyzed per accession. Overall, mean length of stomata ranged between 19.47 microm (diploid) and 27.6 microm (tetraploid), indicating that stomatal length in a tetraploid Malus genotype was 1.4-fold higher than that of a diploid genotype. A positive correlation between genome size and the nucleotypic parameter stomatal length was observed.
苹果属有25至33个物种以及几个亚种。苹果属物种以及同一物种内的克隆体具有不同的倍性水平，因为这些很可能是从不同的树木和（或）不同的地点采集的。近年来，大量苹果属种质资源已在美国国家种质克隆库中收集和保存；然而，这些材料的基因组大小尚未确定。在本研究中，采集了伊利诺伊大学苗圃中生长的100个苹果属物种和杂种的幼龄嫁接树的叶片组织，并立即用于提取细胞核。将苹果和玉米品系W-22的叶片组织作为内标共同切碎，并准备进行流式细胞术分析。苹果细胞核用碘化丙啶（一种嵌入染料）染色，每个样品至少分析8000个细胞核。然后测定苹果细胞核的平均荧光强度。每个样品共进行四次重复。在分析的100个苹果属种质中，鉴定出1个四倍体、3个三倍体和96个二倍体基因型。在所观察到的三种倍性类型之间以及二倍体基因型内均发现了基因组大小的显著差异。四倍体的2C平均值为3.13 pg，三倍体的2C平均值在2.27至2.41 pg之间，而二倍体的2C值在1.44至1.72 pg之间。此外，根据倍性和流式细胞术分析，从10个选定基因型的幼龄树上采集了幼嫩、完全展开叶片的叶印，用于测量核型参数气孔长度。每张载玻片测量10个气孔，每片叶分析3张载玻片，每个种质分析3片叶。总体而言，气孔的平均长度在19.47微米（二倍体）至27.6微米（四倍体）之间，表明四倍体苹果属基因型的气孔长度比二倍体基因型高1.4倍。观察到基因组大小与核型参数气孔长度之间存在正相关。
