家猫中的 Dominant White locus (W) 表现出多效性,显性地导致皮毛缺失色素、不完全显性地导致耳聋和虹膜的浅色。研究者们在白猫家系中进行连锁分析,确定了 KIT 基因为猫的 W locus。在两个家系中对 KIT 基因的分析表明了猫科动物内源性逆转录病毒的插入和进化对纯白和白斑表型的影响。
之前的研究认为,与家猫纯白、白斑性状相关的是同一基因座上的一系列等位基因:W (completely white) > (much spotted) > (little spotted) > (wild-type)。而白手套(white gloving)性状被认为与 KIT 基因连锁。
研究中使用的家猫家系:
Figure 1 Graphic depiction of JHU Pedigree. Pedigree 1 (PI) illustrates matings of white to white cats in the JHU archival colony. Pedigree 2 illustrates pedigree developed to map the Wlocus that is segregating for White coat color. Phenotype of individuals is indicated by color symbol and outline. White symbols denote individuals with a white coat; gray, fully pigmented individuals; half and half symbols (gray/white), white spotted individuals). Hearing capacity is indicated by color outline of the symbol: red outline, deaf; blue, partial hearing; green, normal hearing; black, unknown. Genotypes are depicted below symbol: W, White allele (W, insert of solo LTR; ws, White Spotting allele (insert of full-length FERVelement); w, wild-type (no insertion).可以看到有的纯白个体(白色)、野生型个体(灰色)、白斑个体(一半灰色一半白色);耳聋个体(红色边框)、部分听力丧失(蓝色边框)、听力正常(绿色边框)。黑色边框是听力水平未知。
选择了八个与白色表型有关的候选基因,为它们设计了 STR 引物,并扩增,进行基因型确定。对白色性状、耳聋性状进行连锁分析,用 SUPERLINK 基因进行单标记 LOD (Logarithm of odds) 打分。
结果显示 W 与三个与 KIT 基因连锁的 marker 的连锁很强(LOD非常高,其他基因的都是负数)。于是进一步对 KIT 基因进行分析。
Figure 2 Graphic depiction of feline Chromosome B1 (161.71 Mb-161.62 Mb) (UCSC Genome Browser, September 2011; ICGSC Felis_catus 6.2/felCat5) Assembly. Genomic region of KIT intron1 homologous to murine DNAse hypersensitive site 2 (1) requisite for high-level expression ofKit. Genomic conservation of the region is demonstrated across six mammalian species.对该基因的 21 个外显子的分析没有发现能区别白色和非白色个体的 fixed polymorphism,于是又分析了影响 KIT 调控的区域,发现纯白家猫 KIT 基因的第一个内含子中存在长度为 623 bp 的一段插入。而白斑个体既不是野生型也不是这种 623 bp 插入的突变型。改进了 PCR 方法后,获得了七千多 bp 的 PCR 产物,发现白斑个体是在这一位置有 7125 bp 的全长的猫内源逆转录病毒序列,这段序列与 FERV1 element ERV1-1_FCa-I on chromosome D1 高度相似。
Figure 3 Graphic depiction of retrotransposition of FERV full element and LTR into feline KIT intron 1 in White Spotted and White Dominant individuals; W, White allele; ws, White Spottedallele; w+, wild-type allele.但是不能完全解释白斑家猫中白斑面积表现出的多样性。
Figure 4 Family of domestic cats segregating White Spotting demonstrating difference in degree of White Spotting in individuals inheriting ws allele identical by descent. Squares = males; circles = females. Filled symbols, White Spotted individuals; open symbols, fully pigmented cat. ws, full-length FERV element in KIT;w+, wild-type allele.而对于耳聋性状和浅色虹膜性状,只能得出结论说与 W 位点存在相关性。
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