Development of novel marker loci of rat by detecting variable number of tandem repeats using polymerase chain reaction.

在近交系大鼠s&W中显示变异的遗传标记的鉴定是非常需要的。本研究利用聚合酶链反应（PCR）检测可变数串联重复序列（VNTR），建立了一些标记位点。用PCR方法研究了大鼠白细胞介素-3基因（Il-3）中VNTR的多态性。在28个自交系大鼠中检测到4种类型的扩增片段。分离分析发现Il-3位点与位于大鼠10号染色体上的Gh、RatNGFRR或RatABPG位点存在连锁。结果表明，Il-3基因座位于大鼠第10染色体上，顺序如下：Gh-RatNGFRR-RatABPG-Il-3。大鼠精囊分泌（SVS）IV基因也含有一个VNTR。用PCR扩增该区域，在24株菌株中发现3个等位基因。这种变异以常染色体共显性方式分离。我们将这个位点命名为Svs-4。通过对Svs-4位点与其他位点的连锁分析，发现该位点与Svp-1和一个位点紧密连锁，属于大鼠IV连锁群，因此Svs-4位点属于IV连锁群。在DNA数据库中检索含有二核苷酸重复序列的大鼠微卫星序列。在获得的微卫星序列中，用PCR方法分析了部分序列的大小变化。所有的微卫星序列的大小都是可变的，表明微卫星位点可以作为大鼠连锁分析的有用标记。由此可见，VNTR基因座可作为构建大鼠连锁图谱的有力工具。我们正在利用VNTR位点进行连锁分析，构建大鼠特定染色体的图谱。

Identification of genetic markers exhibiting variations among inbred rat s&W are highly desired. In the present study, some marker loci were established by detecting variable number of tandem repeats(VNTR)using polymerase chain reaction(PCR).A polymorphism of VNTR in rat interleukin-3 gene, Il-3, was investigated by PCR. There were detected four types of the amplified fragments in 28 inbred rat strains. Searegation analysis revealed the linkage between Il-3 locus and Gh, RatNGFRR, or RatABPG loci, which are located on rat chromosome 10. It was thus concluded that the Il-3 locus is located on rat chromosome 10 with following order of loci ; Gh-RatNGFRR-RatABPG-Il-3.The rat seminal vesicle secretion(SVS)IV gene also contains a VNTR. This region was amplified using PCR and tnree alleles were found in 24 strains. This variation segregated as an autosomal codominant manner. We designated this locus as Svs-4. Analysis of linkages between the Svs-4 locus and other loci revealed that this locus is closely linked to Svp-1 and a loci, which belong to rat linkage group IV. Therefor the Svs-4 loci is revealed to belong to the linkage group IV.Microsatellite sequences of rat containing dinucleotide repeats were searched in DNA databases. Among the obtained microsatellites sequences, several sequences were analyzed by PCR to examine size variation in inbred rat strains. All of the microsatellite sequences were variable in size, indicating that the microsatellite loci should be useful marker for linkage analyses of the rat.It was thus concluded that the VNTR loci should be powerful tool for constructing linkage map of the rats. We are now progress in constructing the maps of particular chromosomes of the rats by linkage analyses using the VNTR loci.