关键词: 莲藕根尖细胞, 预处理, 有丝分裂指数, 染色体制片, DAPI染色, 免疫荧光

Abstract: It is difficult to prepare lotus’s（Nelumbo nucifera）root tip chromosome in the cytogenetic study. Therefore，some related factors were studied，such as selecting the proper length of root tip，mitotic index，pretreatment medicament and time，the fixing time，the time of enzyme hydrolysis，and low osmosis in the chromosome preparation from the root tip cells of lotus，and then the immunofluorescence staining was also carried out to validate the chromosome preparation. The results showed that the mitotic index was the highest and morphology was clear when the root tips length was approximately 1.5 cm and the root tips was treated with ice water for 24 hours. The chromosome morphology was solid while root tips was for 1 h fixing with 4% paraformaldehyde. The enzyme hydrolysis was complete，the residue of cytoplasm was little，and the chromosome scattered well while the tips were digesting 30 min with combination of 2.5% cellulase and 2.5% pectinase at 37℃. The chromosome morphology was fine under 30 min low osmosis treatment by 1×PBS. Strong signals were detected after the immunofluorescence staining. This study set up the foundation for the relevant researches on cytogenetic and epigenetic studies of N. nucifera.

Key words: root tip cells of Nelumbo nucifera, pretreatment, mitotic index, chromosome preparations, DAPI staining, immunofluorescence