【作者】 高辉；

【导师】 李旭锋；

【作者基本信息】 四川大学 ， 遗传学， 2003， 硕士

【摘要】 本文对甘蓝型油菜（Brassica napus L.）品种“蜀杂9号”进行了小孢子培养；并对生菜品种“红帆”进行了单细胞培养。结果是：在小孢子培养的试验中，用“蜀杂9号”油菜主花序的长4±0.5mm的花蕾分离出的游离小孢子，使用NLN、MS、B₅培养基（蔗糖浓度使用130g／L），将小孢子的密度调节至10⁴～10⁵个／ml，先在32±1℃中培养48～60h，然后转入25±1℃培养，其中，在NLN培养基中培养的小孢子成胚率最高，并获得白色的愈伤组织。试验结果还表明，NLN培养基要优于MS或B₅培养基；主枝花蕾优于侧枝花蕾；较低的蔗糖浓度（130g／L）要优于较高的蔗糖浓度（170g／L）；高温诱导对小孢子的分裂启动是必须的；而加入活性炭的“液体—固体结合双层培养”方法在小孢子培养过程中没有起到很大的作用。在小孢子培养过程中进行观察，发现小孢子的分裂是小孢子的胞质先变得浓厚，开始逐渐膨大，细胞核也变大，且核的颜色加深；然后在小孢子细胞壁内，细胞核开始分裂，但是细胞壁并没有分裂，只是细胞壁内出现多个核；在正常分裂的小孢子中，有一部分逐渐长大，并最终突破了小孢子的细胞壁，长成为一团细胞团；最后观察到它们逐渐长大成为肉眼可见的白色愈伤组织。 在对生菜（Lactuca sativa）品种“红帆”的细胞培养试验中，进行了不同培养基、细胞密度、激素以及光照等因素对细胞培养成为愈伤组织的影响进行了对比试验。试验结果是：来源于此生菜品种的愈伤组织的单细胞在MS培养基中，并附加0.5mg／L 6-BA和0.1mg／L NAA两种激素，将细胞密度调节至5x104个/ml，置于暗中，25℃进行培养，愈伤生成率最高。比较其培养过程，我们可以看到:MS培养基对生菜细胞培养有着同BS培养基相差无几的效果:而细胞密度则是影响生菜细胞培养的一个重要因素，细胞密度与细胞的生长速度成反比;激素是另一个影响生菜细胞生长的重要因素，在较低浓度6一BA和NAA的共同作用下，细胞生长旺盛;而光照则会降低细胞的生长速度。生菜的单细胞在培养过程中，进行了单细胞一2细胞期一4细胞期一8细胞期一愈伤组织团块的生长方式。更多还原

【Abstract】 In the test, the microspore technique was used to induce haploids from Shuza No.9 of Brassica napus. And at the same time, the suspension cell cultures was studied Hongfan ofLactuca sativa. The microspores were gotten from the buds 4(± 0.5)mm long of the main anthotaxy, then inoculated on the medium of NLN with sucrose of 130g/L. The density of the microspores were adjusted to 104~105/ml, and the microspores were cultured at 32(± 1)℃ in darkness for 48~60h, and then transferred at 25(±1)℃ in darkness. The highest efficiency of induced embryo from microspores of Shuza No.9 was gotten from NLN medium, and the white calluses were found. Other results were gotten from the microspore culture of Shuza No.9: NLN medium was better than MS and 65; the buds of the main anthotaxy excelled the buds from the side anthotaxy; the sucrose of 130g/L overmatched the sucrose of 170g/L in the medium; the initial high temperature was necessary to induce the microspore to start up; the "solid-liquid double-deck" medium added active carbon did not achieve its working. In the process of the microspore culture, the cytoplast of the microspore became dense or the microspore swelled, and the nucleus also got dense and darkened. Then in the cell wall, the microspore divided to several nucleoli, and these nucleoli broke through the cell wall, developed into a dumpling of cells. At last, they became into white calluses.During the cell culture ofLactuca sativa, the effect of some factors on the cell culture was studied such as the type of the basal medium, the density of the cultured cells, the hormone and illumination. The result showed that when the single cell ofhe callus was cultured in the medium of MS+0.5mg/L 6-BA+0.1mg/L NAA, in darkness, at 25 ℃, with 5×104cells/ml, the highest inducement rate of callus was obtained. From the process of the cell culture, some results were acquired: the medium of 85 had the same efficiency with the medium of MS, the density of the cultured cells was an important factor and was inverse pro rata to the growth speed of cells. The low density of hormones (6-BA and NAA) was another important factor to the growth of cells. The illumination decreased the growth speed of cells. In the process, the cell culture of Lactuca sativa experienced single cell-two-cell stage-four-cell stage-eight-cell stage-calluses.更多还原