Optimization of Electroporation Parameters for Immature Embryos of indica Rice (Oryza sativa)

To obtain a suitable condition for electroporation transformation in indica rice, the 10-day-old immature embryos were selected for optimization experiments. The results showed that one pulse at 850 V/cm, 950μF capacitance, 200 μL electroporation buffer with 70 mmol/L sodium glutamate, 100 μg/mL plasmid, 50μg/mL carrier DNA, 20 embryos per cuvette, 0℃ treatment and CC medium were the best parameters, which not only improved the transformation efficiency to 30.89%, but also ameliorated the embryo survival ratio to 95.92%. To further verify the practicability of this condition, the embryos from another indica rice variety and a rice type Ⅱ metallothionein-like gene （OsMT2bL） promotec：mgfp5：：gusA construct were tested, and specific GUS expression on the embryos was visualized by histochemical staining. The results showed that the GUS expression on the embryos activated by the OsMT2bL promoter was mainly concentrated on the apical point of the plumule whereas the expression driven by CaMV35S promoter was distributed on nearly all areas of the electroporated tissues. These results indicated that the optimized embryo electroporation conditions could be used not only in genetic transformation of indica rice but also in assay of gene regulation on embryos.

Analysis of quantitative trait loci underlying the traits related to chlorophyll content of the flag leaf in rice

A population of 117 doubled haploid(DH)lines derived from the cross of Zhaiyeqing 8(indica)6 Jingxi 17(japonica)was employed to map quantitative trait loci(QTL)underlying four physiological traits related to chlorophyll contents of the flag leaf.There were significantly positive correlations among chlorophyll a,chlorophyll b and chlorophyll a+b content.Chlorophyll a/b ratio was significantly negatively correlated with chlorophyll b content.These four traits were normally distributed with transgressive segregation,suggesting that they were controlled by multiple minor genes.A total of 11 QTLs were detected for the four traits and they lay on six chromosomes.Each of them explained 9.2%–19.6%of the phenotypic variations,respectively.Of these,two QTLs controlling chlorophyll a content were mapped on chromosomes 2 and 5;four QTLs underlying chlorophyll b content were mapped on chromosomes 2,3,5 and 9;three QTLs underlying chlorophyll a+b amount were mapped on chromosomes 3,5 and 9;two QTLs underlying chlorophyll a/b ratio were mapped on chromosomes 6 and 11.The intrinsic relationship among the four traits and the practical implication in rice breeding are discussed.

Techniques of cell type-specific transcriptome analysis and applications in researches of sexual plant reproduction

In higher plants, specific cell differentiation and fate decision are controlled by differential gene expression.Cell type-specific transcriptome analysis has become an important tool for investigating cell regulatory mechanisms. Inrecent years, many different techniques have been developed for the isolation of specific cells and the subsequenttranscriptome analysis, and considerable data are available regarding the transcriptional profiles of some specific cells.These cell type-specific transcriptome analyses hold significant promise for elucidating the gene expression linked tocellular identities and functions, and are extraordinarily important for research in functional genomics and systemsbiology aimed toward basic understanding of molecular networks and pathway interactions. Moreover, to reveal thecritical mechanisms about sexual plant reproduction, the gamete and embryo cells have long been treated as goodsubjects for cell-specific transcriptome analysis, and there has been important progress in recent decades. In this review,we summarize current technologies in cell type-specific transcriptome analysis and review the applications of thesetechnologies in research into the mechanisms of sexual reproduction in higher plants.

Cytogenetic comparisons between A and G genomes in Oryza using genomic in situ hybridization

Oryza sativa (一个染色体) 和 O 的 genomic 结构。meyeriana (G 染色体) 比较地在 situ 杂交(GISH ) 用二色的 genomic 被学习。GISH 清楚地能在 O 的染色体之间区别。sativa 和 O。在没有堵住 DNA 的种间的 F1 混血儿的 meyeriana，和合作杂交几乎没被检测。O 的平均有丝分裂的染色体长度。meyeriana 被发现是乘 O 的的 1.69。sativa。染色的 4,6-diamidino-2-phenylindole 的比较出现了 O 的染色体。meyeriana 更广泛地被标记，建议 G 染色体与更重复的序列比被放大一个染色体。在分裂期间原子核， 9-12 多彩石印版中心通常被检测，将近，所有多彩石印版中心组成了 G 染色体特定的 DNA。中心由相应于 G 染色体的染色质压缩形成了的更多和更大的多彩石印版与它的父母相比在混血儿被检测。在 F1 混血儿的 pachytene 期间， A 和 G 的大多数染色体互相，除了 1-2，染色体在他们的手臂的结束配对的触处。在 meiotic 中期我， chromosomal 协会的三种类型，即 O。sativa-O。sativa (A-A ) ， O。sativa-O。meyeriana (A-G ) 和 O。meyeriana-O。meyeriana (G-G ) ，在 F1 混血儿被观察。配对配置的 A-G 染色体包括了 bivalents 和 trivalents。结果向学习染色体组织和 O 的进化提供了一个基础。meyeriana。

Characterization of Interspecific Hybrids Between Oryza sativa L, and Three Wild Rice Species of China by Genomic In Situ Hybridization

In the genus Oryza, interspecific hybrids are useful bridges for transferring the desired genes from wild species to cultivated rice （Oryza sativa L.）. In the present study, hybrids between O. sativa （AA genome） and three Chinese wild rices, namely O. rufipogon （AA genome）, O. officinalis （CC genome）, and O. meyeriana （GG genome）, were produced. Agricultural traits of the F1 hybrids surveyed were intermediate between their parents and appreciably resembled wild rice parents. Except for the O. sativa × O. rufipogon hybrid, the other F1 hybrids were completely sterile. Genomic in situ hybridization （GISH） was used for hybrid verification. Wild rice genomic DNAs were used as probes and cultivated rice DNA was used as a block. With the exception of O. rufipogon chromosomes, this method distinguished the other two wild rice and cultivated rice chromosomes at the stage of mitotic metaphase with different blocking ratios. The results suggest that a more distant phylogenetic relationship exists between O. meyeriana and O. sativa and that O. rufipogon and O. sativa share a high degree of sequence homology. The average mitotic chromosome length of O. officinalis and O. meyeriana was 1.25- and 1.51-fold that of O. sativa, respectively. 4＇,6＇-Diamidino- 2-phenylindole staining showed that the chromosomes of O. officinalis and O. meyeriana harbored more heterochromatin, suggesting that the C and G genomes were amplified with repetitive sequences compared with the A genome. Although chromocenters formed by chromatin compaction were detected with wild rice-specific signals corresponding to the C and G genomes in discrete domains of the F1 hybrid interphase nuclei, the size and number of O. meyeriana chromocenters were bigger and greater than those of O. officinalis. The present results provide an important understanding of the genomic relationships and a tool for the transfer of useful genes from three native wild rice species in China to cultivars.

Heterozygosity of Knob-Associated Tandem Repeats and Knob Instability in Mitotic Chromosomes of Zea （Zea mays L. and Z. diploperennis Iltis Doebley）

Knobs are blocks of heterochromatin present on chromosomes of maize （Zea mays L.） and its relatives that have effects on the frequency of genetic recombination, as well as on chromosome behavior. Knob heterozygosity and instability in six maize inbred lines and one Z. diploperennis Iltis Doebley line were investigated using the fluorescence in situ hybridization （FISH） technique with knob-associated tandem repeats （180 bp and 350 bp （TR- 1）） as probes. Signals of seven heterozygous knobs containing 180- bp repeats and of one heterozygous knob containing TR- 1 were captured in chromosomes of all materials tested according to the results of FISH, which demonstrates that the 180-bp repeat is the main contributor to knob heterozygosity compared with the TR- 1 element. In addition, one target cell with two TR- 1 signals on one homolog of chromosome 2L, which was different from the normal cells in the maize inbred line GB57, was observed, suggesting knob duplication and an instability phenomenon in the maize genome.