摘要
以小麦基因组DNA为模板 ,对ISSR反应程序中的一些重要参数进行摸索和优化试验 ,建立了一套小麦ISSR分析的最优化反应体系及反应程序。即 2 5 μL反应体系中 ,含有 2 0mmol/LMg2 + 、15 0 μmol/LdNTP、2 0 0nmol/L引物、6 0ng模板DNA、2 0UTaqDNA聚合酶。反应程序为第 1个循环基因组DNA经 94℃预变性3min ;4 0个循环为 94℃变性 30s,4 8℃退火 6 0s,72℃延伸 90s ;最后 1个循环完成后 ,在 72℃下延伸 7min。
Inter-Simple Sequence Repeat (ISSR) as a newer technique has been widely used for detection of the DNA development in recent years. Based on the genomic DNA extracted from wheat, some essential factors that might affect the result of ISSR assay were tested and compared. An optimal reaction system suitable for the assay and usage of ISSR in wheat was established, that is, there are 2.0 mmol/L Mg^(2+), 150 μmol/L dNTP, 200 nmol/L primer, 60 ng template DNA, 2.0 U Taq DNA polymerase in a volume of 25 μL amplification reactions system. The reaction program was devised for 3 min at 94℃, 40 cycles of 30 s at 94℃, 60 s at 48℃, 90 s at 72℃, and 7 min at 72℃ in the final cycle.
出处
《河北农业大学学报》
CAS
CSCD
北大核心
2004年第1期10-13,共4页
Journal of Hebei Agricultural University
基金
国家自然科学基金资助项目 (30 170 6 0 2 )
