Comparative efficiency of phosphate-solubilizing bacteria under greenhouse conditions for promoting growth and aloin-A content of Aloe barbadensis

This study was conducted with Aloe barbadensis in order to investigate the efficacy of four phosphate-solubilizing bacteria (PSB), Pseudomonas synxantha 10223, Burkholderia gladioli 10242, Enterobacter hormaechei 10240 and Serratia marcescens 10241 to solubilize Mussorie rock phosphate (MRP) and to evaluate its effects on growth, soluble P content and P uptake compared with control, i.e. uninoculated plants. Pot experiments were conducted in a greenhouse, in soil supplemented with MRP. Each PSB treatment showed different effects on different plant growth parameters. The maximum increase in leaf length (23.7%), total number of leaves (33.33%) and dry rind weight (69.10%) was observed in plants treated with P. synxantha 10223 compared with control. Whereas, maximum increase in root length (23.43%), fresh leaves weight (79.03%), dry gel weight (113.08%) and total gel volume (112.10%), was observed in plants treated with S. marcescens 10241 compared with uninoculated plants. Maximum increase in aloin-A content [114.92% (per g dry gel weight) and 322.32% (per plant dry gel weight)] was observed in plants treated with P. synxantha 10223 compared with control plants. Root colonization by inoculated PSB as estimated by RAPD technique showed that all PSB were able to survive in the rhizosphere of Aloe plants.     

优化反向PCR法分离转基因油菜外源T-DNA侧翼序列的研究(英文)

[目的]优化反向PCR(IPCR)条件分离油菜转基因外源T-DNA的侧翼序列。[方法]以单拷贝的转基因油菜FS4为研究材料,用SDS法和改良CTAB法(包括低温操作、增加酚氯仿的抽提次数以及延长低温沉淀时间等)提取转基因油菜总DNA,并进行酶切、纯化、自连接,再使用普通聚合酶与热启动高保真的ExTaqHS聚合酶进行两轮巢式PCR扩增,进行序列对比。油菜数据库比对搜索采用BBSRCBrassicaDB上WU-BLAST2.0工具,拟南芥数据库比对分析采用TAIR数据库中WU-BLAST2.0工具。为验证FS4转基因中T-DNA插入位点的真实性,根据序列比对分析结果在T-DNA插入位点的左右两端分别设计引物FS4-L与FS4-R。分别采用3对引物pS2/FS4-L、pA2/FS4-R、FS4-L/FS4-R同时对T1代转基因FS4杂合体和非转基因对照植株的基因组进行PCR验证。[结果]两轮巢式PCR扩增得到1个大小为4.0kb的片段,其序列与油菜数据库中BH652424和BZ044084两序列同源性分别高达97.8%和63.4%。根据序列比对结果设计特异引物对进行PCR验证的结果,也证明了FS4转基因油菜中T-DNA的确插入在该位点。[结论]优化后的IPCR条件能成功分离油菜转基因外源T-DNA的侧翼序列。     

油茶随机扩增多态DNA条件的研究

以改进的CTAB法提取油茶嫩叶总DNA,进行随机扩增多态DNA(RAPD)分析,分别测试了Mg2+浓度、引物浓度、dNTP浓度、模板DNA浓度和TaqDNA聚合酶用量对反应结果的影响,确定了油茶RAPD分析的最适反应体系:在25μlPCR反应体积中,含20ng模板DNA,2 5mmol·L-1MgCl2,0 2mmol·L-1dNTP,0 3μmol·L-1引物,1UTaqDNA聚合酶。扩增程序为:94℃预变性180s;94℃变性60s,37℃退火90s,72℃延伸120s,反应40个循环;最后在72℃延伸5min。     

Comparison of the Value of Pulsed-field Gel Electrophoresis,Random Amplified Polymorphic DNA and Amplified rDNA Restriction Analysis for Subtyping Taylorella equigenitalis

Eight strains of Taylorella equigenitalis were identified by a polymerase chain reaction using a primer pair specific to the 16S rDNA of T equigenitalis. These eight strains were chosen because they had previously been shown to represent eight distinct genotypes by pulsed-field gel electrophoresis analysis after separate digestion of the genomic DNA with ApaI or NotI. The eight strains could be classified into six or seven types by random amplified polymorphic DNA analysis using different kinds of primers. Amplified rDNA restriction analysis after separate digestion with five restriction enzymes, including AluI and MboI, of the 1,500 bp fragments of rDNA amplified by polymerase chain reaction did not discriminate the genomic variations among the eight strains of T equigenitalis. Thus, pulsed-field gel electrophoresis was shown to discriminate these eight organisms better than random amplified polymorphic DNA analysis, while amplified rDNA restriction analysis was found to be unsuitable for subtyping T equigenitalis.     

Pod shatter resistance evaluation in cultivars and breeding lines of Brassica napus, B. juncea and Sinapis alba

Pod shatter susceptibility was investigated in Brassica napus germplasm and shatter resistant species of B. juncea and Sinapis alba. The comparisons were made by measuring seed yield in field plots, detached pod rupture energy (RE) and the half‐life of pod‐opening. Pod shatter resistance was significantly greater in B. napus lines derived from interspecific hybridizations of B. napus with B. rapa, B. carinata and B. juncea, than common B. napus cultivars. While these lines exhibited no significant difference in resistance to pod shatter than B. juncea, an entry of S. alba had no yield loss caused by pod shatter. Resistance to pod shatter was characterized in the field as little or no yield loss after full maturity, delayed shattering in time, and stable yield performance under variable climatic conditions during pod maturity. Yield loss caused by pod shatter ranged from a low of 4% for the B. juncea cv. ‘AC Vulcan’ to a high of 61% for the black seeded B. napus line DH12075 in 2‐year field trials after 1 month maturity. Pod shatter resistance was not significantly associated with specific plant and pod morphological traits, except pod length (P = 0.005) in tested materials. Field visual scores of pod shatter through inspections of average pod shatter per plant within plots were highly correlated with plot yield loss. Indoor quantitative evaluations of pod strength using a pendulum machine to measure pod RE and random impact test to measure half‐life of pod‐opening resistance were highly correlated with field yield loss. Multiple evaluations of pod shatter in method and in time after pod maturity are recommended for reliable evaluation of pod shatter resistance.     

Screening RAPD primers for molecular taxonomy and cultivar fingerprinting in the genus Actinidia

Summary Eighty ten-base long arbitrary primers were tested for PCR-based DNA amplification of three species of the genus Actinidia (A. deliciosa the kiwifruit, A. chinensis, and A. kolomikta), with the aim of screening species-specific and genotype-specific markers.Of the 80 primers tested, 30 gave an average of 3.5 bands which were monomorphic within one or two species and absent in the remaining one(s), thus resulting in useful markers for taxonomic and phylogenetic purposes. None of the primers tested produced bands linked to sex. Twenty primers out of the twenty-five selected from a preliminary screening showed high levels of polymorphism, producing two to eleven patterns each from the 13 kiwifruit cultivars examined.We found the Stoffel fragment and the Taq polymerase were both suitable for RAPD analysis, the most noticeable difference being the smaller size of fragments (0.4–1.2 kb) produced by the former in comparison to the latter (1.0–3.4 kb). We tested also three different annealing temperatures (35, 37, and 39° C) and found the intermediate one best for number of amplified bands and reproducibility of results.Abbreviations 2-BE 2-butoxyethanol - CTAB hexadecyltrimethylammonium bromide - MAS Marker-Assisted-Selection - PCR Polymerase Chain Reaction - RAPD Random Amplified Polymorphic DNA - RFLPs Restriction Fragment Length Polymorphisms     

Molecular mapping and characterization of traits controlling fiber quality in cotton

Cotton (Gossypium spp) is the world's leading natural fiber crop. Genetic manipulation continues to play a key role in the improvement of fiber quality properties. By use of DNA-based molecular markers and a polymorphic mapping population derived from an inter specific cross between TM-1 (G. hirsutum) and 3-79 (G. barbadense), thirteen quantitative trait loci (QTLs) controlling fiber quality properties were identified in 3-79, an extra long staple (ELS) cotton. Four QTLs influenced bundle fiber strength, three influenced fiber length, and six influenced fiber fineness. These QTLs were located on different chromosomes or linkage groups and collectively explained 30% to 60%of the total phenotypic variance for each fiber quality property in the F₂ population. The effects and modes of action for the individual QTLs were characterized with 3-79 alleles in TM-1 genetic background. The results indicated more recessive than dominant, with much less additive effect in the gene mode. Transgressive segregation was observed for fiber fineness that could be beneficial to improvement of this trait. Molecular markers linked to fiber quality QTLs would be most effective in marker-assisted selection (MAS) of these recessive alleles in cotton breeding programs. This revised version was published online in August 2006 with corrections to the Cover Date.     

The frequency of natural cross-fertilisation in sequential sowings of winter barley

Summary Estimations of random mating frequency were computed for a series of sequential autumn sowings of populations of winter barley. The estimations were by means of the maximum likelihood scoring method and three varieties, each carrying a recessive genetic marker, provided three independent estimations for each population. High levels of out-crossing were found in sowings made early in September, and a trend towards absence of out-crossing in populations sown in late November was evident. Although fluctuations in this trend could be accounted for by fluctuations in meteorological factors, a more profound underlying effect was evident in that the three marker varieties behaved similarly, though to different degrees. It is postulated that the principle effect of sowing date, as it affects out-crossing, is upon the development of the flowering apex, and that environmental conditions at flowering time merely modify flowering behaviour.     

DNA fingerprinting in soybean (Glycine max (L.) Merrill) with oligonucleotide probes for simple repetitive sequences

Summary Soybean DNA fingerprints were analyzed by digoxigenin-labeled oligonucleotide probes complementary to simple repetitive sequences. The clearest and most polymorphic patterns were obtained with (AAT)₆ as a probe, with which all 47 soybean cultivars tested could be distinguished. However, DNA fingerprints of individuals within cultivars showed the same pattern. Using (CT)₈, (GAA)₅ or (AAGG)₄ as probes, clear polymorphic patterns among cultivars and accessions in the subgenus Soja (Glycine max and Glycine soja) were not observed, while quite different patterns were found in accessions in the subgenus Glycine. The results suggest that G. max and G. soja are closer in their genome structure. DNA fingerprints of reciprocal crosses between cultivars and accessions in the subgenus Soja were similar, and contained bands of both parents. In an F₂ population from these crosses, such bands segregated in a Mendelian fashion.     

Sex-linked SSR markers in hemp

Hemp is a dioecious plant with sex chromosomes X and Y, the male sex being heterogametic. The quality of the fibre depends on the sex type. The sex chromosomes can be characterized by molecular markers. In this report, sex‐linked simple sequence repeat (SSR) markers are described. One SSR marker was polymorphic in both the populations derived from single crosses, two other markers in but one of the two populations. Three alleles were detected for two SSR markers indicating polymorphism not only between X and Y, but also between different X chromosomes. In addition, several sex‐linked RAPD markers were detected in one population. Recombination within the sex chromosomes was observed for nearly all markers.