红莲型水稻不育系与保持系线粒体DNA酶切分析

为研究红莲型水稻细胞质雄性不育的分子基础，我们提取纯化了红莲型水稻丛广４１Ａ和丛广４１Ｂ的线粒体ＤＮＡ，采用限制性内切酶酶切片段长度多态性（ＲＦＬＰ）方法比较了红莲型不育系和保持系线粒体ＤＮＡ酶切图谱，发现两者之间存在一定的差异，部分支持了细胞质雄性不育与线粒体ＤＮＡ有关的结论。     

论“SDNA位移”

提出了“ＤＮＡ复制新假说”，并以此假说分析基因交换、转位、外源基因插入及染色体易位、重复、倒位、缺失等遗传性变异，即“ＳＤＮＡ位移”．认为这些遗传性变异具有相同的发生机理，且发生于ＤＮＡ复制后期．     

牛摩拉氏菌溶血素基因克隆的探讨

应用分子遗传学方法，以光敏生物素标记的ＤＮＡ探讨针进行分子杂交等手段克隆了牛摩拉氏菌溶血素结构基因，并在大肠杆菌中得到表达，其重组子定名为ＭＲ９，分子量为６．４５Ｋｂ，外源片段分子量为３．８Ｋｂ?     

精子介导的报道基因在转基因泥鳅胚胎中的表达

以含ｌａｃＺ基因的质粒ｐＣＨ１１０为外源ＤＮＡ,采用外源ＤＮＡ和精子在保存液里混合保温处理,再进行精,卵的体外受精的方法,观测了精子介导的报道基因在转基因泥鳅胚胎中的表达,４次重复实验均获得了较稳定的实验结果。有９．６％的个体呈现了。ｌａｃＺ基因表达的阳性,对有关实验结果进行了分析和讨论。     

兔桑椹胚胎细胞周期同步化及核移植后的发育能力

进一步改进兔供体桑椹胚Ｇ１期可逆性同步化方法，并比较Ｇ１期核移植胚胎和未同步化核移植胚胎在体内的发育能力。将兔早期１６细胞胚用秋水仙胺和ａｐｈｉｄｉｃｏｌｉｎ处理适当时间，这时期胚胎细胞处于３２细胞期的Ｇ１期。经扫描显微分光光度法测定表明，没有ＤＮＡ合成。经两种药物同步化处理，胚胎正常细胞周期、分裂及移植前发育均不受影响。与用未同步化３２细胞胚细胞核移植相比，用同步在Ｇ１期胚胎细胞进行核移植后，显     

英国发现促癌细胞扩散关键分子

英国癌症研究中心的科学家在近期《细胞学杂志》上报告说 ,他们发现了促使癌细胞扩散的关键分子 ,并利用其制成抗癌疫苗 ,在人体试验中取得了成功。　　英国最初在对胚胎干细胞滋养层和癌细胞进行比较时发现这种名为 5 T4的表面分子。滋养层是胎盘和胎儿之间的一层特殊的胚胎干细胞。由于滋养层与癌细胞同样具备防止人体免疫系统排斥的能力 ,科学家对两者进行了比较 ,发现其中都存在 5 T4分子。他们还发现 ,老鼠胚胎干细胞中的 5 T4分子能够使培养液中的癌细胞不断扩散。另外 ,5 T4分子在癌细胞中表达过量 ,而在成熟的正常细胞中根本不表…     

利用酵母Two-hybrid系统筛选编码产物与p53相作用的人类基因

用酵母ｔｗｏ－ｈｙｂｒｉｄ系统在人胚胎脑组织ｃＤＮＡ表达文库中寻找编码产物与ｐ５３相互作用的基因。在１．５×１０６个转化子中筛选到６８个初级阳性克隆。通过测定转化子第二轮β－半乳糖苷酶活力发现，人胚胎脑组织ｃＤＮＡ表达文库中有一个阳性克隆中的ｃＤＮＡ编码产物与ｐ５３反应结果为明显阳性，说明两个蛋白之间存在较强的相互作用。用热测序法测得这段ｃＤＮＡ的序列通过查询基因数据库发现，这段ｃＤＮＡ编码人的泛肽交联酶，可认为是泛肽交联酶参与ｐ５３在细胞内浓度的调节。     

鸭胚胎血液中PGCs数量及其外源DNA的体内转染

抽取了麻鸭和骡鸭胚胎发育第14－20期的血液，制作血涂片，PAS染色，观察血液中的原生殖细胞的含量，并对各个时期血液中原生殖细胞的含量进行了统计，最高含量都出现在第16期,分别为8．9752％和2．1871％，用In vivo Gene Delivery Systern和pEGFP-N对PGCs进行了外源DNA的体内转染。结果在6只胚胎中的3只中发现GFP报告基因的表达。     

利用酶母Two—hybrid系统筛选编码产物与p53相作用的人类基因

用酵母ｔｗｏ－ｈｙｂｒｉｄ系统在人胚胎脑组织ｃＤＮＡ表达文库中寻找编码产物与ｐ５３相互作用的基因，在１．５×１０＾６个转化子中筛选到６８个初级阳性克隆。通过测定转化子第二轮β－半乳糖苷酶活力发现，人胚胎脑组织ｃＤＮＡ表达文库中有一个阳性克隆中的ｃＤＮＡ编码人的泛肽交联酶，可认为是泛肽交联酶参与ｐ５３在细胞内浓度的调节。     

基因工程与外源DNA导入技术

基因工程是把目的基因与适当的载体相连接形成重组ＤＮＡ, 并引入到适当的寄主细胞中进行复制和表达; 外源ＤＮＡ 导入技术是将供体总ＤＮＡ的片段, 在自花受粉后一定时期内, 使其沿着花粉管通道或其他方法进入胚囊, 转化受精卵或其前后的细胞． 基因工程方法思路清晰、原理清楚、目的明确, 但操作繁琐, 费用高; 外源ＤＮＡ导入技术简便易行, 费用低, 但目的性不强, 工作量大． 如将两种方法结合使用, 可降低费用, 提高整合率． 综述了两种方法的研究进展情况及在实际应用中所取得的成果     

Expression of a microinjected immunoglobulin gene in the spleen of transgenic mice

Transgenic mice were produced by microinjection of a rearranged, functional immunoglobulin kappa gene into fertilized mouse eggs and implantation of the microinjected embryos into foster mothers. Mice that integrated the injected gene were mated and the DNA, RNA and serum kappa chains of their offspring were analysed. The data from offspring of three different transgenic mice indicate that the microinjected gene is expressed in the spleen, but not the liver of mice which inherited the injected gene.     

食蚊鱼卵子发生的组织学观察

食蚊鱼卵子发育过程分为6个时相。第1时相,卵原细胞细胞质较少,核质比较大。第2时相,卵母细胞体积增加迅速,核质比显著减小,细胞质强嗜碱性,滤泡细胞出现。到晚期,有卵黄核及类生长环的出现,这两种结构均和卵黄物质的形成有关。第3时相,卵母细胞嗜碱性减弱,卵黄泡从膜内缘开始出现,逐渐充满细胞质。颗粒层滤泡细胞互相堆积成舌状镶嵌入卵膜,并使放射带凹凸不平,称为镶嵌型滤泡。第4时相,卵母细胞体积增加较快,卵黄泡消失,卵黄物质逐渐充满细胞质,细胞核移至边缘。第5时相,滤泡细胞脱落,成为成熟卵,卵黄物质均匀散布,卵膜极薄。第6时相,未受精卵和滤泡细胞逐渐退化,被吸收。     

Detection of virus-specific sequences in Drosophila melanogaster mutants induced by injection of RSV DNA into early embryos

The injection of purified Rous sarcoma virus (RSV) (Prague strain) into Drosophila melanogaster (Oregon R line) eggs changes the fly phenotype in certain cases, and RSV-specific sequences can be identified in the Drosophila genome (ref. 1 and preceding paper). Here we have used Southern blotting to analyse in greater detail the proviral DNA present in several mutant lines of D. melanogaster produced by microinjection of intact RSV or plasmid DNA containing the viral insert. In certain populations of flies, RSV provirus was found to be incorporated into cellular DNA, and in one mutant family the unintegrated form of plasmid DNA was identified. Generally, the presence of injected genetic material in fly cells correlated with morphological changes in Drosophila.     

Drosophila nurse cells produce a posterior signal required for embryonic segmentation and polarity

The segmental pattern of insect embryos depends on influences from morphogenetic centres near each of the egg poles. In Drosophila, maternal effect mutations are known that impair the normal function of each centre. Injection of wild-type cytoplasm into mutant eggs has revealed that morphogenetic signals localized at the anterior and posterior pole of eggs can be transplanted. We show here that these activities can also be detected during oogenesis. Posterior activity can be recovered at an early stage (stage 10, ref. 5) from the oocyte-nurse cell complex, but anterior activity can only be detected in the mature oocytes (stage 14). We conclude that the bicoid-dependent anterior signal, although produced by the nurse cells, does not become active before it is localized to the anterior egg pole, whereas posterior activity can be detected in the nurse cells before, and therefore independently of, its localization to the posterior egg pole.     

Application of a nuclear localization signal gene in transgene mice

Efficient gene transfer by cytoplasm co-injection will offer a powerful means for transgenic animals. Using co-injection in cytoplasm, two independent gene constructs, including bovine α-s1-casein-hG-CSF and a mammal expression vector expressing a nuclear localization signal (mNLS), were introduced into fertilized mouse eggs. The target gene construct was docked into host nucleus probably by the nuclear localization signal. Transgene mice have been obtained at 58% (29/50) of integration ratio. Expression level of the positive transgene mice was detected by Western blotting. Maximal expression of human G-CSF was estimated about 540 mg/L of milk. The expression ratio was up to 75% (9/12). The results here have important practical implications for the generation of mammary gland bioreactors and other transgene studies. Co-injection of a target gene with an expression vector of a mammal nuclear localization signal by cytoplasm appears to be a useful, efficient and easy strategy for generating transgenic animals, which may be able to substitute the routine method of pronucleus-injection of fertilized eggs.     

灭幼脲对云斑天牛的不育作用及核酸含量的影响

用灭幼脲处理的补充营养寄主植物饲养云斑天牛Ｂａｔｏｃｅｒａｈｏｒｓｆｉｅｌｄｉ（Ｈｏｐｅ）成虫，对子代卵的孵化有明显抑制作用，生化分析表明：灭幼脲可以促进雌虫卵巢组织ＤＮＡ，ＲＮＡ及一日龄卵ＤＮＡ的合成；对一日龄卵内ＲＮＡ含量影响不显著，对发育末期胚胎的ＤＮＡ，ＲＮＡ有明显抑制作用。结合灭幼脲在世代间垂直传递途径，讨论了灭幼脲对云斑天牛成虫不育作用的机理。     

MAPK regulates cell cycle progression in pig oocytes and fertilized eggs

Oocytes collected from prepubertal gilt ovaries were matured in vitro (IVM), and fertilized in vitro (IVF) or electrically activated. Phosphorylation of mitogenactivated protein kinase (MAPK) was detected with SDS-PAGE and Western blotting, and translocation of ERK₂ was observed with immunofluorescent cytochemistry. We found that the quantity of MAPK kept unchanged during oocyte maturation. There was no phosphorylated MAPK in porcine oocytes at the germinal vesicle (GV) stage; a little MAPK was phosphorylated at 20 h of IVM; a high level phosphorylation was detected at 30 h, while MAPK phosphorylation decreased at 36 h; and then MAPK phosphorylation increased again to the peak level from 40 to 60 h. ERK2 translocated from the peripheral cytoplasm to inner cytoplasm and nuclear area during oocyte maturation. There was nearly no phosphorylated MAPK at 18 and 20 h of electrically activated oocytes, but phosphorylation increased at 22 h. There was no phosphorylated MAPK at 12 h of IVF, while phosphorylation resumed at 16 h. These results suggest that MAPK may play an important regulatory role in MI-MII transition, pronucleus formation and the initiation of the first mitosis in pig eggs.     

用原位杂交技术对玉米中期染色体中rRNA的研究

本文以玉米１７ＳｒＲＮＡ的ｃＤＮＡ作探针，采用缺口平移技术，用Ｂｉｏ－ｄＵＴＰ对该探针进行生物素标记。在Ｋ４Ｍ树脂包埋的玉米根尖超薄切片上进行原位杂交。杂交后，在电镜下通过与１０ｎｍ金颗粒相连的亲合素对杂交子进行检测，首次直接证实了除核仁，核质和细胞质外，ｒＲＮＡ在中期染色体上的存在。其分布特点是随机的，均匀的，并在数量上明显高于细胞质。     

拟松材线虫个体发育研究

【目的】探索拟松材线虫(Bursaphelenchus mucronatus)的繁殖能力,并对其胚胎发育过程中经历的重要阶段和卵的形态变化进行研究,了解胚胎发育和完成整个生活史所需时间,为进一步研究其生长发育并进行有效防控提供参考。【方法】分别挑取3组180条拟松材线虫雌成虫,观察记录雌虫在25 ℃条件下的产卵情况,每隔2 h统计每组线虫的累积产卵量,直至卵的数量基本不再增加。挑取尚未产卵的拟松材线虫雌虫于载玻片上,待其产卵后,将卵置于蔡司体视显微镜下观察。连续观察胚胎的发育过程并使用照片记录不同发育阶段胚胎的形态变化,记录卵发育至不同阶段所需时间。挑取约200个刚产下的拟松材线虫卵,在25 ℃条件下发育24 h后每隔4 h统计其总孵化率,直至孵化数不再增加,设置3组重复。将刚孵化的2龄幼虫接种于灰葡萄孢(Botrytis cinerea)上,分为3组,每组设置3个重复,分别在接种1、2、3 d后使用贝尔曼漏斗法收集线虫,计算混合龄线虫中每龄期线虫所占比例,计算拟松材线虫胚后发育及完成整个生活史所需时间。【结果】① 在拟松材线虫产卵能力方面,0~10 h拟松材线虫产卵总量增长较快,16 h后产卵量逐渐趋于稳定,28 h内雌虫平均累积产卵12粒/条。② 拟松材线虫的胚胎发育过程主要经历以下几个关键阶段:单胞期、双胞期、3胞期、4胞期、5胞期、8胞期、16胞期、囊胚期、利马豆期、蝌蚪期、蠕虫期、1龄幼虫(J1),至孵化为2龄幼虫(J2)时结束。③ 在胚胎发育前期,第1次卵裂发生的位置存在两种情况,即卵的1/2和1/3处。双胞发育至5胞时也存在两种不同的发育方式,一种是双胞不移动直接分裂成3胞并列排列,另外一种是细胞进行移动,3胞呈三角形排列。通过观察30个卵的第1次卵裂和100个卵双胞的发育过程发现,这些不同的发育方式均是普遍存在的。④ 在25 ℃条件下,拟松材线虫卵的累积孵化率随时间增加而增加,在32 h时达到最高(93.31%),随后逐渐趋于稳定。⑤ 在25 ℃条件下记录了拟松材线虫卵从单胞发育至各个阶段的时间,完成整个胚胎发育过程需要约28 h。2龄幼虫接种于灰葡萄孢3 d后即可获得新的2龄幼虫,因此拟松材线虫完成整个生活史只需要3 d。【结论】对拟松材线虫卵从单胞阶段直至孵化的整个胚胎发育过程进行观察发现,拟松材线虫完成胚胎发育大约需要28 h,完成整个生活史需要3 d。对拟松材线虫产卵能力和卵的孵化率进行统计,收集拟松材线虫卵和2龄幼虫的最佳时间分别为16 h和36 h。拟松材线虫胚胎发育前期,在第1次卵裂期以及由双胞期发育至5胞期的两个过程中均存在两种与同属线虫不同的发育方式。这种现象还有待进一步研究。     

Discovery of phosphatized gastrula fossils from the Doushantuo Formation, Weng’an, Guizhou Province, China

Globular fossils from Doushantuo phosphorites at the Weng’an area are for the first time identified as a gastrulation stage of phosphatized embryos. They are got from the fossiliferous remains after acetic acid maceration. The fossils are found together with formerly reported animal resting eggs and embryos of the earlier cleavage stage. The oblate-shaped fossils with the same size as those reported embryos and invaginate at the middle part into the embryos, show the characteristics of the late blastula to the early gastrula stage of the embryo development. This discovery convinces the existence of animal embryos at Doushantuo age and offers new facts for the studying of the affinity of related fossils, which are still controversial at present.