Supporting research on making linkage maps of marker, RFLP and RAPD loci in genome of sugi, Cryptomeria japonica D.Don

杉杉（Sugi, Cryptomeria japonica D.Don）自古以来就是日本重要的林业树种。单倍体染色体数目为11，除两条染色体具有卫星缩窄或次级缩窄外，其他表型特征相似。本项目首先研究了在存在或不存在致死因子的情况下，同工酶、RFLP和RAPD等标记位点之间重组值的估计方法。通过观察到的分离比与每个位点的预期分离比的偏差大小来检验致死位点的存在。采用LOD评分评价模型中致死因子的适合度。用该方法对同工酶进行分离，分别与Lap、G6p、Dia-3连锁。在Kumotooshi和Okinoyama sugi杂交的3代后代中，共构建了164个位点，包括128个RFLP、33个RAPD、2个同工酶和1个矮子，共13个连锁群，总图谱距离为1190 cM。在13个连锁群中，有8个连锁群存在分离比偏差的聚集位点。在6个连锁群中检测到1个或2个致死基因，发现有了致死基因，图谱距离比无致死基因估计的要长。通过对单倍体巨生植物的分析，绘制了另一张包含175个RAPD位点的图谱。共有14个连锁群，总长度2212.3 cM。采用冷处理和荧光原位杂交技术对单倍体染色体进行了鉴定。尝试改进方法，用激光束切割后恢复了染色体碎片。

Sugi, Cryptomeria japonica D.Don has been a important forestry species in Japan sice old times. It has haploid chromosome number of 11 with similar phenotypic features except to two chromosomes which have a satelite or secondary constriction.In this project, firstly, studies were made on the methods for estimatin of recombination values between marker loci including isozyme, RFLP and RAPD loci with or without the presence of a lethal factor. Existence of a lethal locus was examined by the magnitude of deviation of the observed segregation ratio from expected one for each locus separately. The fitness of the model involving a lethal factor were evaluated by LOD scores. This method was applied for isozyme segregation and it was found a lethal gene linked to Lap, G6p, Dia-3 respectively.In a three generation progeny derived from a cross of Kumotooshi and Okinoyama sugi, 164 loci including 128 RFLP,33 RAPD,2 isozyme and 1 dwarf were put in a linkage map with 13 linkage groups with total map distance of 1,190 cM.Out of 13 linkage groups, there were 8 linkage groups which had clustered loci with deviated segregation ratios. One or two lethal genes were detected in 6 linkage groups, and it was found that with the lethal genes, map distances were elongated than those estimated by free of lethal gene. Another map in which 175 RAPD loci were placed, was made by analysis of haploid megagametophytes. It had 14 linkage groups with 2,212.3 cM in total.Discrimination of haploid chromosome was tried by clod treatment and fluorescence in situ hybridization. It was tried to improve the methods and chromosome fragments were recovered after cutting with laser beam.