Fundamental study for citrus chromosome mapping

本研究的主要目的是为柑橘核型分析开辟新的染色体制备方法，这是基因染色体定位的重要内容；二是尝试用这种新方法进行柑橘核型分析，并为柑橘染色体鉴定开辟有用的染色体显带方法。离体培养植株根尖的最佳酶处理为2%纤维素酶+ 1%果胶酶120 min或3%纤维素酶+ 2%果胶酶90min，果园幼树叶尖的最佳酶处理为3%纤维素酶+ 1%果胶酶90min。每条染色体中最长的一条染色体长度达到4-10倍，是已报道的最长染色体长度的2-5倍以上。本研究的主要染色体数为：柑桔和柑桔的2n=18，‘托沙-邦丹’和‘穗-邦丹’的2n=20-21，柑桔的2n=19，福图纳拉的2n=20，柚子的2n=19-20。对Poncirus和orange的核型分析表明，Poncirus和orange均有2对常着心染色体和7对亚常着心染色体，其中Poncirus染色体可鉴定6对，orange染色体可鉴定5对。G显带法和N显带法均不适用。C带法显示出独特的带型，但在本研究中重复性很低。CMA/DAPI染色在Tosa-Buntan染色体上显示稳定的独特条带。染色体有2带、1带和无带三种类型。

Major purpose of this study is to develop new chromosome preparation method for citrus karyotype analysis that is important of gene chromosome mapping, and the second is to try citrus karyotype analysis using this new method and to develop useful chromosome banding method for identification of citrus chromosome.1.Optimum enzymatic treatment was 120 min.incubation of 2% cellulase + 1% pectolyase or 90 min.incubation of 3% cellurase + 2% pectolyase for root tip of in vitro cultured plantlet, and 90min.incubation of 3% cellurase + 1% pectolyase for young leaf tip from orchard tree. The chromosome length of the longest one in each chromosome setreached 4-10mum that was over 2-5 times of the longest chromosome length which had been reported.2.The chromosome numbers of 2n=18 in Poncirus and orange, 2n=20-21 in 'Tosa-Buntan' and 'Suisho-Buntan', 2n=19 in satsuma mandarin, 2n=20 in Fortunella and 2n=19-20 in yuzu were major in this investigation. The karyotyape analysis of Poncirus and orange indicated that both had 2 pairs of metacentric and 7 pairs of submetacentric chromosome, and 6 paires of Poncirus chromosome and 5 pairs of orange chromosome were possible to be identified.3.G banding and N banding method was not available for citrus chromosome. C banding method showed unique banding pattern but repeatability was very low in the present study. CMA/DAPI stain showed stable unique band in 'Tosa-Buntan' chromosome. There were 3 types of chromosome that having 2 bands, 1band and no band.