Department of Biotechnology, Indian Institute of Technology, Guwahati, Assam, India.
Plant Signal Behav. 2011 Jul;6(7):974-81. doi: 10.4161/psb.6.7.15503.
Present report is the first direct evidence of azadirachtin production in androgenic haploid cultures of Azadirachta indica, a woody medicinal tree. Anther cultures at early-late-uninucleate stage of microspores were established on MS medium with BAP (5 μM), 2,4-D (1 μM) and NAA (1 μM) containing 12% sucrose. The calli, induced, were further multiplied on 2,4-D and Kinetin media. Shoots, differentiated on BAP (2.2 μM) + NAA (0.05 μM) medium, were elongated on MS + BAP (0.5 μM) and multiplied on MS + BAP (1 μM) + CH (250 mg/l). Thereafter, the shoots were rooted on ¼ MS + IBA (0.5 μM). Cytological analysis of the calli and regenerants have confirmed their haploid status with the chromosome number as 2n = x = 12. The haploid cell lines and leaves from in vitro grown plantlets were analyzed for azadirachtin by RP-HPLC and mass spectroscopy. Maximum azadirachtin (728.41 μg/g DW) was detected in calli supporting best shoot proliferation while least (49 μg/g DW) was observed in an undifferentiated line from maintenance medium. This study has brought us a step closer to develop genetically pure lines that could serve as new and attractive alternative ways of homogeneous controlled production of high value compounds, round the year, independent of geographical and climatic barrier.
本报告首次直接证明印楝在雄性单倍体培养物中产生印楝素，印楝是一种木本药用树。在含有 BAP（5 μM）、2,4-D（1 μM）和 NAA（1 μM）的 MS 培养基上建立小孢子早晚期单核期的花药培养。诱导的愈伤组织在 2,4-D 和激动素培养基上进一步繁殖。在 BAP（2.2 μM）+NAA（0.05 μM）培养基上分化的芽在 MS+BAP（0.5 μM）上伸长，并在 MS+BAP（1 μM）+CH（250 mg/l）上繁殖。之后，芽在 ¼ MS+IBA（0.5 μM）上生根。对愈伤组织和再生体的细胞学分析证实了它们的单倍体状态，染色体数为 2n = x = 12。通过反相高效液相色谱法和质谱法分析来自体外生长的植物的单倍体细胞系和叶片中的印楝素。在支持最佳芽增殖的愈伤组织中检测到最大量的印楝素（728.41 μg/g DW），而在维持培养基中的未分化系中观察到最小量（49 μg/g DW）。这项研究使我们更接近开发遗传纯系，这些纯系可以作为全年生产高价值化合物的新的有吸引力的替代方法，不受地理和气候障碍的影响。
