Hunger H D, Speer A, Flachmeier C, Hanke R, Behrendt G, Coutelle C
Academy of Sciences of the GDR, Department of Human Molecular Genetics, Berlin.
Anal Biochem. 1987 Aug 15;165(1):45-55. doi: 10.1016/0003-2697(87)90199-0.
Conditions for the optimal use of cyanuric chloride-activated (CCA) paper in Southern transfer hybridization experiments of genomic DNA were investigated. They depend critically on pH and ionic strength during transfer and on the composition of the hybridization solution. Simplified hybridization conditions using a SSC/dextran sulfate system at 65 degrees C without sodium dodecyl sulfate and the complex Denhardt's solution are applied. CCA paper allows repeated use in hybridization experiments. Under optimized conditions CCA paper allows a more sensitive detection of single-copy gene sequences in the subpicogram range than do nylon membranes. Application of these transfer and hybridization conditions with our newly developed CCA paper to carrier determination and prediction of the healthy male haplotype demonstrates its usefulness for prenatal counseling of a Duchenne muscular dystrophy family.
研究了在基因组DNA的Southern转移杂交实验中，氰尿酰氯活化（CCA）纸的最佳使用条件。这些条件在很大程度上取决于转移过程中的pH值和离子强度以及杂交溶液的组成。采用了在65℃下使用不含十二烷基硫酸钠的SSC/硫酸葡聚糖系统和复杂的Denhardt溶液的简化杂交条件。CCA纸可在杂交实验中重复使用。在优化条件下，与尼龙膜相比，CCA纸能够更灵敏地检测亚皮克范围内的单拷贝基因序列。将这些转移和杂交条件与我们新开发的CCA纸应用于携带者检测和健康男性单倍型预测，证明了其在杜氏肌营养不良症家庭产前咨询中的实用性。
