Department of Genetics, Katholieke Universiteit Nijmegen, Toernooiveld, 6525 ED Nijmegen, The Netherlands.
EMBO J. 1983;2(10):1741-6. doi: 10.1002/j.1460-2075.1983.tb01651.x.
We microdissected a Y chromosomal lampbrush loop pair from primary spermatocyte nuclei of Drosophila hydei and cloned the DNA directly at the microscale. Four of the 12 recombinant DNA clones recovered display in situ hybridization to mitotic metaphase Y chromosomes, preferentially in the chromosomal region identified as the origin of the lampbrush loop pair. All clones, however, also hybridize to autosomal and X chromosomal loci in polytene chromosomes. Y chromosomal DNA sequences of D. hydei again prove to be members of different families of repeated sequences distributed throughout the genome. These microcloning experiments, which were carried out under very unfavourable experimental conditions (low DNA content of the lampbrush loops in the presence of large amounts of RNA) prove that almost any chromosomal structure detected by light microscopy is directly accessible to molecular cloning experiments by micromethods.
我们从小果蝇精母细胞细胞核的 Y 染色体灯刷环对中进行显微解剖,并直接在微观尺度上克隆 DNA。回收的 12 个重组 DNA 克隆中有 4 个显示有丝分裂中期 Y 染色体的原位杂交,优先在被鉴定为灯刷环对起源的染色体区域。然而,所有克隆也与多线染色体中的常染色体和 X 染色体位置杂交。小果蝇的 Y 染色体 DNA 序列再次证明是分布在整个基因组中的不同重复序列家族的成员。这些微克隆实验是在非常不利的实验条件下进行的(在存在大量 RNA 的情况下,灯刷环的 DNA 含量低),证明了通过微量方法,几乎任何通过光学显微镜检测到的染色体结构都可以直接进行分子克隆实验。