Kikuchi Y, Mori K, Suzuki S, Yamaguchi K, Mizuno S
Department of Agricultural Chemistry, Faculty of Agriculture, Tohoku University, Sendai, Japan.
Gene. 1992 Jan 15;110(2):151-8. doi: 10.1016/0378-1119(92)90642-3.
Two overlapping genomic clones containing the fibroin light-chain (Fib-L)-encoding gene (Fib-L) were obtained from the cosmid library of the silkworm, Bombyx mori J-139, by hybridization with the Fib-L cDNA clone. Sequencing of the 14.6-kb region revealed that Fib-L was 13,472 bp long containing seven exons, and that the gene contained a large first intron which occupied about 60% of the gene. Comparison of restriction patterns of the J-139 Fib-L with those of eight other B. mori breeds producing normal-level fibroin demonstrated that considerable restriction-fragment length polymorphisms were present in regions containing the first intron and the 3'-flanking sequence. However, sizes of the Fib-L mRNA and the Fib-L polypeptide were very similar among the nine breeds tested, suggesting that the exon sequences and the splice signals were all well conserved. 5'-Flanking regions of Fib-L and the fibroin heavy-chain (Fib-H)-encoding gene (Fib-H) compared in this study contained three 18-30-bp sequences of high similarity and many 8-10-bp common elements, six of which coincided with the binding sites of homeodomain proteins. Gel retardation assays with the nuclear extracts of the posterior and middle silk glands suggested that protein factors present in the posterior silk-gland nuclei could bind to a set of those common upstream elements.
通过与丝心蛋白轻链(Fib-L)cDNA克隆杂交,从家蚕Bombyx mori J-139的黏粒文库中获得了两个重叠的基因组克隆,其中包含编码Fib-L的基因(Fib-L)。对14.6 kb区域的测序表明,Fib-L长13472 bp,包含7个外显子,并且该基因含有一个大的第一内含子,约占该基因的60%。将J-139 Fib-L的限制性酶切图谱与其他8个产生正常水平丝心蛋白的家蚕品种的图谱进行比较,结果表明在包含第一内含子和3'侧翼序列的区域存在相当多的限制性片段长度多态性。然而,在所测试的9个品种中,Fib-L mRNA和Fib-L多肽的大小非常相似,这表明外显子序列和剪接信号都得到了很好的保守。本研究中比较的Fib-L和丝心蛋白重链(Fib-H)编码基因(Fib-H)的5'侧翼区域包含三个18 - 30 bp的高度相似序列和许多8 - 10 bp的共同元件,其中六个与同源结构域蛋白的结合位点一致。用后部和中部丝腺的核提取物进行凝胶阻滞分析表明,后部丝腺细胞核中存在的蛋白质因子可以与一组这些共同的上游元件结合。