Study for citrus chromosome mapping

为了制作柑橘染色体物理图谱，需要确定以母本营养组织为材料，染色体较长，便于核型分析和染色体鉴定的染色体制备方法。在染色体制备方法中，利用3-5mm的幼叶，用0.3%纤维素酶+0.2%胸胶酶溶液在37℃下处理120-180min，将悬浮细胞滴在玻片上，可以制备较长的染色体。2 ． 根据CMA带型将柑橘染色体分为5种类型。即在染色体上，CMA（+）区域位于双臂末端和近端（a型），位于单臂末端和近端（b型），位于双臂末端（c型），位于单臂末端（d型），没有CMA（+）区域（e型）。依赖CMA带型的柑橘染色体组成为：佛手2B+8D+8E，曲1C+8D+9E， unshiu 1A+1C+8D+8E, ‘Washington’脐橙2B+2C+7D+7E， ‘Duncan’柚子2A+1B+1C+6D+8D， ‘Tosa-Buntan’ 1A+1B+5C+2D+9E, ' suiso - buntan ' 3A+3C+3D+9E, ' baniso -okan ' 2A+1B+3C+3D+9E, 'Banpeiyu ‘ 1A+2B+2C+4D+8E+1A或B， ’ tritriate橙子4B+8D+6E。对‘Nankan N0.20’unshiu染色体，用Dig标记的‘Nankan N0.20’DNA和Biotin标记的‘Tosa-Buntan’DNA进行基因组原位杂交（GISH），并用抗Dig罗丹明和亲和素FITC对染色体进行染色。将‘南坎N0.20’8条d型染色体分为黄色大信号1号染色体、黄色小信号1号染色体、橙色大信号3号染色体、橙色小信号1号染色体和红色小信号2号染色体。此外，通过吉氏染色将8条e型染色体分为2长、2中、4短染色体。

For making physical map of citrus chromosomes , chromosome preparation method in which vegetative tissue of mother plant use as a material and chromosomes are relatively long and useful for karyotype analysis , and identification of chromosomes should be confirmed.1. In chromosome preparation method, using young leaves of 3-5mm, 120-180min treatment of 0.3% cellurase +0.2% pectolyase solution at 37℃ and dropping suspended cells to slide-glass were useful for relatively long chromosome prepared.2 . Citrus chromosomes were classtfied to 5 types based on CMA banding patterns . That is , CMA(+) regions in terminal of both arms and a proximal (type-A) , in terminal of one arm and a proximal (type-B) , in terminal of both arms (type-C), in terminal of one arm (type-D) and no CMA(+) region (type-E) on chromosome.3. Citrus chromosome compositions depended on CMA banding pattern were 2B+8D+8E in fingered citron, 1C+8D+9E in koji, 1A+1C+8D+8E in unshiu, 2B+2C+7D+7E in 'Washington' navel orange, 2A+1B+1C+6D+8D in 'Duncan' grapefruit, 1A+1B+5C+2D+9E in 'Tosa-Buntan' , 3A+3C+3D+9E in 'Suisho-Buntan' , 2A+1B+3C+3D+9E in ' Ban-okan ' , 1A+2B+2C+4D+8E+1A or B in 'Banpeiyu ' and 4B+8D+6E in triforiate orange.4. For 'Nankan N0.20' unshiu chromosomes, genomic in situ hybridization (GISH) was performed using Dig labeled 'Nankan N0.20 ' DNA and Biotin labeled 'Tosa-Buntan ' DNA, and chromosomes were stained with anti-Dig rhodamine and avidin FITC. Eight type-D chromosomes of ' Nankan N0.20 ' were classified to I chromosome with large yellow signal, I with small yellow signal, 3 with large orange signal, I with small orange signal and 2 with small red signal. Furthermore in 'Nankan N0.20 ' chromosomes, 8 type-E chromosomes were classified to 2 long, 2 medium and 4 short chromosomes by Giemsa staining.