Genome rearrangement and nitrogen fixation in Anabaena blocked by inactivation of xisA gene

Two genome rearrangements involving 11- and 55-kilobase DNA elements occur during the terminal differentiation of an Anabaena photosynthetic vegetative cell into a nitrogen-fixing heterocyst. The xisA gene, located on the nifD 11-kilobase DNA element, was inactivated by recombination between the chromosome and a copy of the xisA gene that was mutated by inserting an antibiotic gene cassette. Site-directed inactivation of the Anabaena xisA gene blocked rearrangement of the 11-kilobase element and nitrogen fixation, but did not affect rearrangement of the 55-kilobase element, heterocyst differentiation, or heterocyst pattern formation.     

Genetic transformation of Drosophila with transposable element vectors

Exogenous DNA sequences were introduced into the Drosophila germ line. A rosy transposon (ry1), constructed by inserting a chromosomal DNA fragment containing the wild-type rosy gene into a P transposable element, transformed germ line cells in 20 to 50 percent of the injected rosy mutant embryos. Transformants contained one or two copies of chromosomally integrated, intact ry1 that were stably inherited in subsequent generations. These transformed flies had wild-type eye color indicating that the visible genetic defect in the host strain could be fully and permanently corrected by the transferred gene. To demonstrate the generality of this approach, a DNA segment that does not confer a recognizable phenotype on recipients was also transferred into germ line chromosomes.     

Heritable somatic excision of a Drosophila transposon

A mutation in the white gene of Drosophila mauritiana that results from insertion of the transposable element mariner is genetically unstable in both germ cells and somatic cells. Somatic instability is indicated by the occurrence of animals having mosaic eyes with patches of pigmented cells on a peach-colored background. Normally uncommon, the frequency of mosaicism is so greatly enhanced in a particular mutant strain that virtually every animal in the strain is an eye-color mosaic. The molecular basis of the mosaicism is the excision of the mariner element from its location in the DNA of the white gene in somatic cells. The phenomenon results from a single dominant genetic factor located in chromosome 3. Genetic control over the excision of transposable elements may play a role in determining the persistence of transposable elements in the genome.     

Stable integration and expression of a bacterial gene in the mosquito Anopheles gambiae

Foreign DNA was successfully introduced into the germline of the African mosquito vector of malaria Anopheles gambiae. Stable integration of genes into the germlines of insects had been achieved previously only in Drosophila melanogaster and related species and required the use of the P element transposon. In these experiments with Anopheles gambiae, the plasmid pUChsneo was used, which contains the selectable marker neo gene flanked by P element inverted repeats. Mosquitoes injected with this plasmid were screened for resistance to the neomycin analog G-418. A single event of plasmid insertion was recovered. Integration appears to be stable and, thus far, resistance to G-418 has been expressed for eight generations. The transformation event appears to be independent of P.     

A trans-acting product needed for P factor transposition in Drosophila

A transposable genetic element of the P family in Drosophila melanogaster was found to be unstable in the presence of other P elements but stable in their absence. A sensitive assay for P transpositional activity is provided by the snw allele, a defective P insert in the singed bristle locus which becomes hypermutable only in the presence of complete elements. This measure of activity was highly correlated with a type of female sterility normally associated with P activity. There was no cross-reactivity with transposase from another hybrid dysgenesis-causing element (the I factor).     

Correction

In the Research News article "A ;mitey' theory for gene jumping" (6 Sept., p. 1093), Jean Marx erred when she wrote that the groups of Marilyn A. Houck and Margaret G. Kidwell (Reports, 6 Sept., p. 1125) focused on Proctolaelaps regalis as a possible carrier of P elements among Drosophila species by a "process of elimination." Viruses were not screened for P elements during the study, as implied, nor were any other organisms. Rather, the work was initiated by acarologist Houck (then on the faculty of the University of Arizona), who was aware of P element history from conversations with the Kidwell lab. Houck and Ken Peterson, a postdoc in the Kidwell lab, then initiated the Southern blot analysis that pointed to the presence of P element DNA in P. regalis, and Jonathan Clark, a postdoc at Arizona's Center for Insect Science, significantly extended the work through the application of polymerase chain reaction. Their data do not exclude the possibility of viral participation in the system, however.     

Xotch, the Xenopus homolog of Drosophila notch

During the development of a vertebrate embryo, cell fate is determined by inductive signals passing between neighboring tissues. Such determinative interactions have been difficult to characterize fully without knowledge of the molecular mechanisms involved. Mutations of Drosophila and the nematode Caenorhabditis elegans have been isolated that define a family of related gene products involved in similar types of cellular inductions. One of these genes, the Notch gene from Drosophila, is involved with cell fate choices in the neurogenic region of the blastoderm, in the developing nervous system, and in the eye-antennal imaginal disc. Complementary DNA clones were isolated from Xenopus embryos with Notch DNA in order to investigate whether cell-cell interactions in vertebrate embryos also depend on Notch-like molecules. This approach identified a Xenopus molecule, Xotch, which is remarkably similar to Drosophila Notch in both structure and developmental expression.     

Regulated splicing of the Drosophila P transposable element third intron in vitro: somatic repression

In eukaryotic cells alternative splicing of messenger RNA precursors (pre-mRNA's) is a means of regulating gene expression. Although a number of the components that participate in regulating some alternative splicing events have been identified by molecular genetic procedures, the elucidation of the biochemical mechanisms governing alternative splicing requires in vitro reaction systems. The tissue specificity of P element transposition in Drosophila depends on the germline restriction of pre-mRNA splicing of the P element third intron (IVS3). Drosophila P element IVS3 pre-mRNA substrates were spliced accurately in vitro in heterologous human cell extracts but not in Drosophila somatic cell splicing extracts. Components in Drosophila somatic cell extracts that specifically inhibited IVS3 splicing in vitro were detected by a complementation assay. Biochemical assays for Drosophila RNA binding proteins were then used to detect a 97-kilodalton protein that interacts specifically with 5' exon sequences previously implicated in the control of IVS3 splicing in vivo. Inhibition of IVS3 splicing in vitro could be correlated with binding of the 97-kD protein to 5' exon sequences, suggesting that one aspect of IVS3 tissue-specific splicing involves somatic repression by specific RNA-protein interactions.     

Rescue of the Drosophila phototransduction mutation trp by germline transformation

Phototransduction is the process by which light-stimulated photoreceptor cells of the visual system send electrical signals to the nervous system. Many of the steps that follow the initial event in phototransduction, absorption of light by rhodopsin, are ill-defined. The fruitfly, Drosophila melanogaster, provides a means to dissect phototransduction genetically. Mutations such as transient receptor potential (trp) affect intermediate steps in phototransduction. In order to facilitate molecular studies of phototransduction, the trp gene was isolated and its identity was confirmed by complementing the mutant trpCM allele of the trp gene by P-element mediated germline transformation of a 7.1-kilobase DNA fragment. Expression of the trp gene begins late in pupal development and appears to be limited to the eyes and ocelli.     

Transmission of integrated sea urchin histone genes by nuclear transplantation in Xenopus laevis

Sea urchin histone genes contained in a recombinant plasmid pSp102 were microinjected into the cytoplasm of fertilized eggs of Xenopus laevis. By the late blastula stage, plasmid DNA sequences were detected comigrating with the high molecular weight cellular DNA (greater than 48 kilobases). Analysis of the DNA from injected embryos digested with various restriction endonuclease demonstrated that the injected DNA was integrated into the frog genome. Clones of embryos containing the pSp102 DNA sequences were produced by means of nuclear transplantation. Individuals of the same clone contain the pSp102 sequences integrated into similar chromosomal locations. These sites vary between different clones.     

Cloning and expression of a Xenopus embryonic gap junction protein

Gap junctions in the early amphibian embryo may play a fundamental role in the regulation of differentiation by mediating the cell-to-cell transfer of chemical signals. A complementary DNA encoding a gap junction present in Xenopus oocytes and early embryos has now been cloned and sequenced. This protein sequence is homologous to the well-characterized gap junction structural proteins rat connexin32 and connexin43. RNA blot analysis of total Xenopus oocyte RNA showed hybridization to a single 1.6-kilobase band. This messenger RNA is abundant in oocytes, decreases to levels below the sensitivity of our assay by stage 15 (18 hours), and is not detectable in RNA from a number of adult organs. To confirm that the oocyte cDNA encodes a gap junction channel, the protein was over expressed in Xenopus oocytes by injection of RNA synthesized in vitro. Pairs of RNA-injected oocytes formed many more time- and voltage-sensitive cell-cell channels than water-injected pairs.     

CNS and hypoderm regulatory elements of the Drosophila melanogaster dopa decarboxylase gene

Expression of the dopa decarboxylase gene (Ddc) is regulated in a tissue- and developmental stage-specific manner throughout the life cycle of the fruit fly, Drosophila melanogaster. Essential Ddc regulatory elements lie within 208 base pairs upstream from the RNA start point. Functional elements within this 5' flanking region were mapped by deletion analysis, which assayed expression in vivo after germline integration via P element vectors. One of the elements is essential for expression in both the larval and adult central nervous system, and at least two other elements are necessary for quantitatively normal expression in the hypoderm. Within each of the intervals that have regulatory effects are found sequence elements conserved between the Ddc genes of two distantly related species of flies. On the basis of this correlation, regulatory functions for these sequence elements can be postulated.     

Introduction of human DNA into mouse eggs by injection of dissected chromosome fragments

A procedure has been developed for introducing exogenous DNA into mouse eggs by injection of chromosome fragments. Chromosome fragments were dissected from human metaphase spreads and microinjected into the pronuclei of fertilized mouse eggs. Many of the injected eggs subsequently exhibited normal pre- and postimplantation development. Embryos obtained from eggs injected with centromeric fragments retained human centromeric DNA as demonstrated by in situ hybridization analysis. From eggs injected with noncentromeric fragments, a mouse was obtained whose tail tissue exhibited the presence of human DNA. This procedure should facilitate incorporation of very large (more than 10 megabases) DNA fragments into cells and embryos without the need for cloned sequences.     

一个新的水稻TE的克隆及其与CACTA转座子结构的比较

对水稻受稻瘟病菌诱导的新因子Rim2的基因组结构进行了研究.以Rim2 cDNA片段作探针, 筛选水稻BAC文库并对其亚克隆, 获得了一个基因组DNA克隆Rim2-569.序列分析表明, Rim2-569序列具备Class 2 转座子的基本结构特征.它两端具有完整的末端颠倒重复(TIRs), 若干正向和反向的亚末端重复 (STRs) , 以及插入位点3 bp的同向重复.它TIRs上的保守序列CACTG有别于以往报道的CACTA转座子.该因子包含一个开读框, 其预测蛋白与CACTA转座子编码的TNP2、TNPD等转座酶有低程度的同源性.该因子没有能够编码类似TNP1/TNPA 的DNA结合蛋白的开读框.Southern杂交显示, Rim2因子在多个不同起源的水稻品种上广泛存在,连同检索结果,证明该家族具有很多拷贝.上述特点表明, 该因子属于一个与CACTA转座子有一定差异的新的转座子大家族.     

欧李花粉微形态与矿质营养代谢及其相关性研究

以实生欧李群体为试材,采用扫描电镜观测方法,研究同一生境条件下欧李花粉超微形态结构以及花粉和成熟期果实中矿质元素含量变化及其相关关系,以期为欧李矿质营养代谢机制和钙高效基因型早期筛选提供理论参考.结果表明:欧李花粉形状为长球形或超长球形,极面观为三裂圆形,具三孔沟,按NPC系统分类属于N3P4C5型:花粉外壁纹饰呈穿孔纹饰、条纹-穿孔纹饰和条纹状纹饰三种类型;不同株系在花粉形状、大小、表面纹饰、极轴长(P)、赤道轴长(E)、P/E以及花粉和果实中矿质元素含量等生理形态指标变异较大.相关分析结果表明,花粉中Ca、Mg元素与花粉极轴和赤道轴呈显著或极显著正相关,花粉中K元素与赤道轴和P/E分别呈极显著负相关和显著正相关,花粉中Fe元素与赤道轴呈显著正相关;果实中Ca、K元素与极轴呈显著正相关;花粉和果实中Mn元素与P/E均呈显著负相关.     

金环胡蜂食用虫态矿质元素含量分析

以微波消解法处理金环胡蜂(Vespa mandarinia Smith)4种食用虫态即幼虫、初化蛹、老熟蛹和成虫,采用电感耦合等离子体发射光谱法(ICP-OES)对这4种食用虫态样品中的K、P、S、Ca、Mg、Fe、Cu、Co、Mn、Se、Zn、Ba、Al、Cd等14种矿质元素成分进行了分析。结果表明:金环胡蜂各食用虫态中的K、P、S、Ca、Mg、Fe、Zn和Se等元素含量特别丰富,具有较高的食用价值;有害元素中Cd的含量为0.09～0.36μg/g,尚在国家食品污染物限量范围内,而Al的含量96.78～320.52μg/g,偏高。