Senger G, Jones T A, Fidlerová H, Sanséau P, Trowsdale J, Duff M, Sheer D
Human Cytogenetics Laboratory, Imperial Cancer Research Fund, London, UK.
Hum Mol Genet. 1994 Aug;3(8):1275-80. doi: 10.1093/hmg/3.8.1275.
Free DNA was prepared from routinely harvested and fixed cells for high resolution FISH mapping using either a sodium hydroxide/ethanol mixture or 70% formamide. Hybridization signals from cosmid probes appeared as extended lines. The average length of signals on DNA prepared with sodium hydroxide was significantly greater than with formamide. A set of overlapping cosmids from the HLA class II region was used to determine how precisely the actual overlap or gap between probes can be calculated from the measured overlap or gap between their signals. Lengths of the probe signals and their known kilobase lengths were used as an internal ruler. The mean values calculated from the measured length from 30 or more signals for each probe pair showed remarkable conformity with the known kilobase lengths of their overlaps and gaps. Immediately adjacent probes could also be ordered on the released DNA. These simple procedures dramatically increase the speed with which relationships between probes can be determined during contig construction.
游离DNA是从常规收获并固定的细胞中制备的，用于使用氢氧化钠/乙醇混合物或70%甲酰胺进行高分辨率荧光原位杂交（FISH）图谱分析。黏粒探针的杂交信号呈现为延伸的线条。用氢氧化钠制备的DNA上信号的平均长度显著长于用甲酰胺制备的DNA上信号的平均长度。一组来自HLA II类区域的重叠黏粒用于确定从探针信号之间测量的重叠或间隙能多精确地计算出探针之间实际的重叠或间隙。探针信号的长度及其已知的千碱基长度被用作内部标尺。对于每对探针，从30个或更多信号的测量长度计算出的平均值与它们重叠和间隙的已知千碱基长度显示出显著的一致性。紧邻的探针也可以在释放的DNA上排序。这些简单的程序极大地提高了在构建重叠群过程中确定探针之间关系的速度。
