5—氮胞苷介导的转基因cry1Ab的阶段复活及其对水稻的生物学效应

用不同浓度的和5－氮胞苷溶液配合不同时间处理了发生的在转录水平的转基因cry1Ab沉默水稻。结果表明，5－氮胞苷处理能使8％－30％转基因水稻植株的转基因cry1Ab恢复表达活性，复活性基因表达的Cry1Ab蛋白含量可达0.147％，45mg/L5-氮胞苷处理1d可使转基因cry1Ab的复活率及Cry1Ab蛋白含量达最高。同时，5－氮胞苷处理使转基因水稻植株出现矮化，分蘖数、单株粒重下降，穗长减小及开花期延迟等发育异常现象。5－氮胞苷作为一种弱诱变剂在克服转基因沉默及创造具优良农艺性状的转基因植物中具有一定的应用价值。     

The tryptophan-rich domain of puroindoline is directly associated with the starch granule surface as judged by tryptic shaving and mass spectrometry

The starch granule surface is a frontline of microbial attack and defence, operating in the background of normal starch granule metabolism. Puroindoline, a wheat protein which binds starch granule surfaces, contains a unique tryptophan-rich domain likely responsible for this property, though direct evidence is lacking. To test puroindoline’s tight association, prime starch granule extracts were water-washed 8 or 20 times and residual puroindoline removed using a solution of 50% isopropanol/50 mM NaCl. We found that this solvent was consistent in the amount of protein extracted from wheat flour and washed starch, regardless of initial protein content. Relative quantification of puroindoline following water-washing was performed using dot blot. Washing more than 8 times did not further reduce puroindoline content of starch granules suggesting a strong association with the starch granule surface. To identify the tryptophan-rich domain tightly associated with the starch granule surface, a combination of in situ tryptic digestion and mass spectrometry was used. Following digestion and water-washing, 50% isopropanol/50 mM NaCl was used to remove tightly-associated peptides for identification by mass spectrometry. Using this method, we identified the tryptophan-rich domain of puroindoline directly bound to the starch granule surface of wheat.     

Commercially Produced Rice and Maize Starches Contain Nonhost Proteins,as Shown by Mass Spectrometry

To evaluate the presence of contaminating nonhost proteins in commercially prepared rice and maize starch samples, we initiated a direct sequencing mass spectrometric proteomics survey. We discovered nonhost proteins from a variety of species, including Phytophthora cinnamomi, Homarus americanus, and Ovis aries. Our documentation of H. americanus proteins in these starch samples may have food safety implications with regard to shellfish allergies. We hypothesize that these proteins were introduced to the starch samples via process wash water used in the milling and deproteination steps in the commercial preparation of the starches analyzed. The introduction of nonhost proteins during commercial processing of starch samples that are used routinely in analytical studies indicates that these studies are using impure materials. Therefore, further study and documentation of the starch samples is required to ensure that all components of the samples are properly catalogued.     

Inheritance and field performance of transgenic Korean Bt rice lines resistant to rice yellow stem borer

Transgenic Korean rice plants containing the cry1Ab gene were developed for resistance against yellow stem borer (Scirpophaga incertulas, YSB). More than 100 independent transgenic lines from three Korean varieties (P-I, P-II and P-III) were generated. The amount of Cry1Ab in transgenic T₀ plants was as high as 2.88% of total soluble proteins. These levels were sufficient to cause 100% mortality of YSB larvae. The majority of T₁ transgenic lines originated from the varieties P-I and P-II followed a Mendelian fashion of segregation. Deviation from the expected segregation ratio was observed in a small number of the transgenic lines of P-I and P-II origins. However, this deviation was primarily observed in the P-III originated lines. Segregation analysis of the T₁ generation indicated that 1–3 copies of the cry1Ab gene were integrated into the genome of the majority of the transgenic lines originating from varieties P-I and P-II. Stunted and semi-fertile mutants were observed in some transgenic lines. These aberrations were either independent or closely linked to the introduced cry1Ab gene loci in different transgenic lines. Reduction in GUS expression levels and loss of toxicity against YSB larvae were found in some transgenic lines. The transgenic T₃ and T₄ lines causing 100% mortality of third instar YSB larvae in the lab were completely protected in the field. Analysis of important yield components on nine selected transgenic lines indicated that stem length, panicle length, grain number per panicle, and seed setting rates were reduced in transgenic plants compared to those in non-transgenic parental rice lines. Number of panicles per cluster, however, was significantly higher in transgenic plants. The numerical value of the average yield was in general greater in the controls than in all the transgenic lines, indicating some ‘yield drag’. Since some selected lines were highly resistant to the YSB with good yielding potential, they offer effective potential for use in insect resistance management programs.     

cry1Ab 基因在转基因"克螟稻"后代中的遗传稳定性及表达

以不同世代的转基因“克螟稻”后代为实验材料,研究了ｃｒｙ１Ａｂ基因在不同世代间的遗传稳定性及表达特性。结果表明：ｃｒｙ１Ａｂ基因在“克螟稻”后代中不仅能通过有性世代稳定传递而且能在有性世代交替中保持表达的稳定性,同时ｃｒｙ１Ａｂ基因在“克螟稻”后代中表达表现出了一定的时空特性,主茎、一次分蘖和二次分蘖各部位Ｃｒｙ１Ａｂ蛋白含量均不同、但均以茎、叶、叶鞘中表达量较高,根中表达量则相对较低,其中以成熟     

5-氮胞苷介导的转基因crylAb的阶段复活及其对水稻的生物学效应

用不同浓度的5-氮胞苷溶液配合不同时间处理了发生在转录水平的转基因cry1Ab沉默水稻。结果表明，5-氮胞苷处理能使8%～30%转基因水稻植株的转基因cry1Ab恢复表达活性，复活转基因表达的Cry1Ab蛋白含量最高可达0.147%，45mg/L5-氮胞苷处理1d可使转基因cry1Ab的复活率及Cry1Ab蛋白含量达最高。同时，5-氮胞苷处理使转基因水稻植株出现矮化，分蘖数、单株粒重下降，穗长减小及开花期延迟等发育异常现象。5-氮胞苷作为一种弱诱变剂在克服转基因沉默及创造具优良农艺性状的转基因植物中具有一定的应用价值。     

Agronomic and morphological characterization of Agrobacterium-transformed Bt rice plants

The agronomic and morphological characteristics of Agrobacterium-transformed rice plants carrying the synthetic cry1Ab or cry1Ac gene were investigated. Tremendous variations in plant height, seed fertility, grain size and other traits were seen in 80 T₁ lines, derived from 80 T₀ plants of 9 rice varieties. On average, about 33% T₁ lines had either morphological or agronomic variant plants. Most of the variations in T₁ plants had no significant correlation with transgene insertion and were proved heritable to their progenies. Genetic analysis in T₃ or T₄ generations showed some simple mutations such as chlorophyll deficiency and stunted plants were independent of transgene insertion and seemed to be controlled by a pair of single genes. However, in two independent transgenic progenies of Xiushui 11, all plants homozygous for transgenes showed dwarfism while all hemizygous and null segregants had normal plant heights. Two advanced homozygous Bt lines, KMD1 and KMD2, were developed from these two progenies. Comparison of the agronomic traits of KMD1 and KMD2 with their parent displayed marked differences among them in terms of seedling growth, tillering ability, yield components and yield potential. The genetic variation observed was generally not linked to the transgene locus and was ascribed to somaclonal variation, but other causes might also exist in particular cases. The results are discussed in the context of choosing appropriate transformation methodology for rice breeding programs. This revised version was published online in August 2006 with corrections to the Cover Date.     

Bt转基因水稻“克螟稻”杂交后代二化螟抗性研究初报

利用室内叶片快速鉴定法结合田间抗性调查, 研究了粳型Bt转基因水稻“克螟稻”与籼、 粳稻杂交后代对二化螟(Chilo suppressalis)的抗性。 结果显示, 在与籼稻和粳稻的杂交后代(F1、 F2和BC1)中, 都出现了对二化螟表现高抗的植株。 用离体叶片法测定, 4个组合120株杂种F1和88株GUS阳性(GUS+)BC1株与30株克螟稻亲本相似,