Palmer M J, Buck D A, Frelinger J A, Wettstein P J
J Immunol. 1985 Aug;135(2):1450-5.
Until recently, the analysis of Class II genes linked to the rat major histocompatibility complex, RT1, has been confined to serologic and electrophoretic analysis of their gene products. To obtain a more definitive estimate of the number and relative polymorphism of RT1 Class II sequences, we performed Southern blot analysis of rat genomic DNA employing human cDNA probes specific for Class II heavy and light chain genes. Southern blots of EcoRI and BamHI digests of genomic DNA from ten inbred strains, expressing eight RT1 haplotypes, were hybridized with the human DQ beta or DR beta cDNA that are homologous to Class II light chain sequences. Four to eight bands were observed to hybridize with the light chain cDNA: band sizes ranged from 2.5 to 28 kb. Restriction fragment patterns were polymorphic; the only identical patterns observed were those associated with RT1 haplotypes with identical RT1.B regions. The number and size of bands hybridizing with DQ beta and DR beta suggested a minimum of four light chain sequences in each haplotype. Southern blots of BamHI and EcoRI digests of genomic DNA from the same strains were hybridized with a DR alpha cDNA that is homologous to Class II heavy chain sequences. All RT1 haplotypes expressed either a 10.0-kb or 13.0-kb band when digested with BamHI, and either a 17-kb or 3.7-kb band when digested with EcoRI. Considerably less polymorphism was detected with the DR alpha probe; this observation is consistent with previously reported limited protein polymorphism of the rat equivalent of the I-E alpha subunit. The size and number of bands hybridizing with the DR alpha probe suggests a minimum of two heavy chain sequences. These observations suggest that the RT1 complex includes more Class II sequences than have been observed in serologic and electrophoretic analyses of Class II gene products. Furthermore, the level of polymorphism of RT1 Class II sequences appears to be comparable with mouse and human Class II sequences.
直到最近，与大鼠主要组织相容性复合体RT1相关的Ⅱ类基因分析还局限于对其基因产物的血清学和电泳分析。为了更确切地估计RT1Ⅱ类序列的数量和相对多态性，我们用人Ⅱ类重链和轻链基因特异性的cDNA探针，对大鼠基因组DNA进行了Southern印迹分析。用表达8种RT1单倍型的10个近交系大鼠的基因组DNA进行EcoRI和BamHI酶切，所得的Southern印迹与与人Ⅱ类轻链序列同源的人DQβ或DRβ cDNA杂交。观察到有4至8条带与轻链cDNA杂交：条带大小在2.5至28 kb之间。限制性片段模式具有多态性；观察到的唯一相同模式是与具有相同RT1.B区域的RT1单倍型相关的模式。与DQβ和DRβ杂交的条带数量和大小表明每个单倍型至少有4个轻链序列。用相同品系的基因组DNA进行BamHI和EcoRI酶切，所得的Southern印迹与与人Ⅱ类重链序列同源的DRα cDNA杂交。所有RT1单倍型在用BamHI酶切时都表达一条10.0 kb或13.0 kb的条带，在用EcoRI酶切时都表达一条17 kb或3.7 kb 的条带。用DRα探针检测到的多态性要少得多；这一观察结果与先前报道的大鼠I-Eα亚基等效物的有限蛋白质多态性一致。与DRα探针杂交的条带大小和数量表明至少有2个重链序列。这些观察结果表明，RT1复合体包含的Ⅱ类序列比在Ⅱ类基因产物的血清学和电泳分析中观察到的要多。此外，RT1Ⅱ类序列的多态性水平似乎与小鼠和人类的Ⅱ类序列相当。
