【摘要】 采用切口平移法标记外源物种(黑麦)总基因组DNA作为探针,用受体物种总基因组DNA(中国春小麦)作遮盖,对小麦1B/1R易位系中外源染色体的黑麦易位片段进行检测。对影响基因组原位杂交技术实验效果的因素进行了分析。通过实验认为:细胞分裂相多、染色体分散良好、染色体长度适中的高质量染色体制片是取得理想实验效果的基础;保证在整个实验过程中染色体一直附着在载玻片上不丢失是实验进行的支撑条件;探针DNA与封阻DNA的比例是取得理想实验效果的关键。对杂交信号过多或过少或者无杂交信号的技术环节进行了分析,并提出了解决办法。实验过程中应严格控制每一步骤,才能获得理想的原位杂交实验效果。更多还原

【Abstract】 The genome in situ hybridization(GISH) is an ideal technology of the direct detection of target DNA in the nucleus in the presence and distribution,which particularly facilitates in the identification exogenous chromosomes or chromosome segments of additional lines,substitution line,translocation lines genetic material and genome composition of allopolyploid species.By nick translation labeled exogenous species(rye) total genomic DNA as a probe,with receptor species total genomic DNA(China Spring Wheat) as a cover,exogenous chromosomes of rye translocation fragments was detected in 1B/1R wheat translocation lines,the key experimental technology links of GISH technology were introduced,and influencing factors were analyzed.Experiments show that(1) the high quality chromosome preparation is the foundation of obtaining the ideal experimental effect with phase cell,chromosome,chromosome length of well dispersed with moderate;(2) that chromosome has been attached to the slide,which may not lose the supporting conditions during the whole experiment process;(3) the ratio of probe DNA to blocking DNA is key of obtaining the ideal effect;(4) technical links to hybridization signals too little or too much or no hybridization signal were analyzed.Every step should be strictly controlled in order to obtain the desired effect in situ hybridization experimental process.更多还原