Methodological and reporting standards for quality-of-life data eligible for European Society for Medical Oncology-Magnitude of Clinical Benefit Scale (ESMO-MCBS) credit.

BACKGROUND: The European Society for Medical Oncology-Magnitude of Clinical Benefit Scale (ESMO-MCBS) has been developed to grade clinical benefit of cancer therapies. Improvement in quality of life (QoL) is considered relevant, especially in the non-curative setting. This is reflected by an upgrade of the preliminary ESMO-MCBS score if QoL is improved compared to the control arm or a downgrade if an improvement in progression-free survival is not paralleled by an improvement in QoL or overall survival. Given the importance of QoL for the final score, a need to ensure the robustness of QoL data was recognised. DESIGN: A checklist was created based on existing guidelines for QoL research. Field testing was carried out using clinical trials that either received an adjustment of the preliminary ESMO-MCBS score based on QoL or had QoL as the primary endpoint. Several rounds of revision and re-testing of the checklist were undertaken until a final consensus was reached. RESULTS: The final checklist consists of four items and can be applied if three prerequisites are met: (i) QoL is at least a secondary endpoint, (ii) evidence of reliability and validity of the instrument is provided, and (iii) a statistically and clinically significant improvement in QoL is observed. The four items on the checklist pertain to the (i) hypothesis, (ii) compliance and missing data, (iii) presentation of the results, and (iv) statistical and clinical relevance. Field testing revealed that a clear QoL hypothesis and correction for multiple testing were mostly lacking, while the main statistical method was always described. CONCLUSIONS: Implementation of the ESMO-MCBS QoL checklist will facilitate objective and transparent decision making on QoL data within the ESMO-MCBS scoring process. Trials published until 1 January 2025 will have to meet the prerequisites and at least two items for crediting QoL benefit in the final ESMO-MCBS score. Trials published thereafter will have to meet all four items.

Tuscany Sangiovese grape juice imparts cardioprotection by regulating gene expression of cardioprotective C-type natriuretic peptide.

PURPOSE: A regular intake of red grape juice has cardioprotective properties, but its role on the modulation of natriuretic peptides (NPs), in particular of C-type NP (CNP), has not yet been proven. The aims were to evaluate: (1) in vivo the effects of long-term intake of Tuscany Sangiovese grape juice (SGJ) on the NPs system in a mouse model of myocardial infarction (MI); (2) in vitro the response to SGJ small RNAs of murine MCEC-1 under physiological and ischemic condition; (3) the activation of CNP/NPR-B/NPR-C in healthy human subjects after 7 days' SGJ regular intake. METHODS: (1) C57BL/6J male and female mice (n = 33) were randomly subdivided into: SHAM (n = 7), MI (n = 15) and MI fed for 4 weeks with a normal chow supplemented with Tuscany SGJ (25% vol/vol, 200 µl/per day) (MI + SGJ, n = 11). Echocardiography and histological analyses were performed. Myocardial NPs transcriptional profile was investigated by Real-Time PCR. (2) MCEC-1 were treated for 24 h with a pool of SGJ small RNAs and cell viability under 24 h exposure to H2O2 was evaluated by MTT assay. (3) Human blood samples were collected from seven subjects before and after the 7 days' intake of Tuscany SGJ. NPs and miRNA transcriptional profile were investigated by Real-Time PCR in MCEC-1 and human blood. RESULTS: Our experimental data, obtained in a multimodal pipeline, suggest that the long-term intake of SGJ promotes an adaptive response of the myocardium to the ischemic microenvironment through the modulation of the cardiac CNP/NPR-B/NPR-C system. CONCLUSIONS: Our results open new avenue in the development of functional foods aimed at enhancing cardioprotection of infarcted hearts through action on the myocardial epigenome.

Exploring the genetic diversity of Ethiopian grass pea (Lathyrus sativus L.) using EST-SSR markers.

Expressed sequence tags (ESTs) in public databases and cross-species transferable markers are considered to be a cost-effective means for developing sequence-based markers for less-studied species. In this study, EST-simple sequence repeat (SSR) markers developed from Lathyrus sativus L. EST sequences and cross-transferable EST-SSRs derived from Medicago truncatula L. were utilized to investigate the genetic diversity among grass pea populations from Ethiopia. A total of 45 alleles were detected using eleven EST-SSRs with an average of four alleles per locus. The average polymorphism information content for all primers was 0.416. The average gene diversity was 0.477, ranging from 0.205 for marker Ls942 to 0.804 for MtBA32F05. F(ST) values estimated by analysis of molecular variance were 0.01, 0.15, and 0.84 for among regions, among accessions and within accessions respectively, indicating that most of the variation (84%) resides within accessions. Model-based cluster analysis grouped the accessions into three clusters, grouping accessions irrespective of their collection regions. Among the regions, high levels of diversity were observed in Gojam, Gonder, Shewa and Welo regions, with Gonder region showing a higher number of different alleles. From breeding and conservation aspects, conducting a close study on a specific population would be advisable for genetic improvement in the crop, and it would be appropriate if future collection and conservation plans give due attention to under-represented regions. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s11032-011-9662-y) contains supplementary material, which is available to authorized users.

Identification of putative stage-specific grapevine berry biomarkers and omics data integration into networks.

The analysis of grapevine (Vitis vinifera) berries at the transcriptomic, proteomic, and metabolomic levels can provide great insight into the molecular events underlying berry development and postharvest drying (withering). However, the large and very different data sets produced by such investigations are difficult to integrate. Here, we report the identification of putative stage-specific biomarkers for berry development and withering and, to our knowledge, the first integrated systems-level study of these processes. Transcriptomic, proteomic, and metabolomic data were integrated using two different strategies, one hypothesis free and the other hypothesis driven. A multistep hypothesis-free approach was applied to data from four developmental stages and three withering intervals, with integration achieved using a hierarchical clustering strategy based on the multivariate bidirectional orthogonal projections to latent structures technique. This identified stage-specific functional networks of linked transcripts, proteins, and metabolites, providing important insights into the key molecular processes that determine the quality characteristics of wine. The hypothesis-driven approach was used to integrate data from three withering intervals, starting with subdata sets of transcripts, proteins, and metabolites. We identified transcripts and proteins that were modulated during withering as well as specific classes of metabolites that accumulated at the same time and used these to select subdata sets of variables. The multivariate bidirectional orthogonal projections to latent structures technique was then used to integrate the subdata sets, identifying variables representing selected molecular processes that take place specifically during berry withering. The impact of this holistic approach on our knowledge of grapevine berry development and withering is discussed.

Correction: High throughput approaches reveal splicing of primary microRNA transcripts and tissue specific expression of mature microRNAs in Vitis vinifera.

UNLABELLED: The version of this article published in BMC Genomics 2009, 10:558, contains data in Table 1 which are now known to be unreliable, and an illustration, in Figure 1, of unusual miRNA processing events predicted by these unreliable data. In this full-length correction, new data replace those found to be unreliable, leading to a more straightforward interpretation without altering the principle conclusions of the study. Table 1 and associated methods have been corrected, Figure 1 deleted, supplementary file 1 added, and modifications made to the sections "Deep sequencing of small RNAs from grapevine leaf tissue" and "Microarray analysis of miRNA expression". The editors and authors regret the inconvenience caused to readers by premature publication of the original paper. BACKGROUND: MicroRNAs are short (~21 base) single stranded RNAs that, in plants, are generally coded by specific genes and cleaved specifically from hairpin precursors. MicroRNAs are critical for the regulation of multiple developmental, stress related and other physiological processes in plants. The recent annotation of the genome of the grapevine (Vitis vinifera L.) allowed the identification of many putative conserved microRNA precursors, grouped into multiple gene families. RESULTS: Here we use oligonucleotide arrays to provide the first indication that many of these microRNAs show differential expression patterns between tissues and during the maturation of fruit in the grapevine. Furthermore we demonstrate that whole transcriptome sequencing and deep-sequencing of small RNA fractions can be used both to identify which microRNA precursors are expressed in different tissues and to estimate genomic coordinates and patterns of splicing and alternative splicing for many primary miRNA transcripts. CONCLUSIONS: Our results show that many microRNAs are differentially expressed in different tissues and during fruit maturation in the grapevine. Furthermore, the demonstration that whole transcriptome sequencing can be used to identify candidate splicing events and approximate primary microRNA transcript coordinates represents a significant step towards the large-scale elucidation of mechanisms regulating the expression of microRNAs at the transcriptional and post-transcriptional levels.

High throughput approaches reveal splicing of primary microRNA transcripts and tissue specific expression of mature microRNAs in Vitis vinifera.

BACKGROUND: MicroRNAs are short (approximately 21 base) single stranded RNAs that, in plants, are generally coded by specific genes and cleaved specifically from hairpin precursors. MicroRNAs are critical for the regulation of multiple developmental, stress related and other physiological processes in plants. The recent annotation of the genome of the grapevine (Vitis vinifera L.) allowed the identification of many putative conserved microRNA precursors, grouped into multiple gene families. RESULTS: Here we use oligonucleotide arrays to provide the first indication that many of these microRNAs show differential expression patterns between tissues and during the maturation of fruit in the grapevine. Furthermore we demonstrate that whole transcriptome sequencing and deep-sequencing of small RNA fractions can be used both to identify which microRNA precursors are expressed in different tissues and to estimate genomic coordinates and patterns of splicing and alternative splicing for many primary miRNA transcripts. CONCLUSION: Our results show that many microRNAs are differentially expressed in different tissues and during fruit maturation in the grapevine. Furthermore, the demonstration that whole transcriptome sequencing can be used to identify candidate splicing events and approximate primary microRNA transcript coordinates represents a significant step towards the large-scale elucidation of mechanisms regulating the expression of microRNAs at the transcriptional and post-transcriptional levels.

Addressing drought tolerance in maize by transcriptional profiling and mapping.

In order to unravel the genetic architecture underlying plant response to drought, we adopted an integrated approach, combining transcript profiling and quantitative trait loci (QTL) mapping. In fact, improving plant tolerance to water stress is an important, but, at the same time, a difficult task, since plant tolerance is the result of many complex mechanisms acting at different levels of plant organization, and its genetic basis is largely unknown. The phenotypic data, concerning yield components and flowering time, of a population of 142 maize Recombinant Inbred Lines (RILs), grown under well watered conditions or under water stress, were submitted to linkage analysis to detect drought-tolerance QTLs. Thirty genomic regions containing 50 significant QTLs distributed on nine chromosomes were identified. At the same time, a customized targeted oligoarray was used to monitor the expression levels of 1,000 genes, representative of the immature maize kernel transcriptome. Using this DNA array we compared transcripts from 10 days after pollination kernels of two susceptible and two drought tolerant genotypes (extracted from our RILs) grown under control and water stress field conditions. Two hundred and fifty-two genes were significantly affected by stress in at least one genotype. From a set of these, 49 new molecular markers were developed. By mapping most of them and by in silico mapping other regulated sequences, 88 differentially expressed genes were localized onto our linkage map, which, added to the existing 186 markers, brought their total number on the map to 274. Twenty-two of the 88 differentially expressed genes mapped in the same chromosomal segments harbouring QTLs for tolerance, thus representing candidate genes for further functional studies.

Identification of quantitative trait Loci for resistance to southern leaf blight and days to anthesis in two maize recombinant inbred line populations.

The genetic architecture underlying resistance in maize to southern leaf blight (SLB) caused by Cochliobolus heterostrophus race O is not well understood. The objective of this study was to identify loci contributing to SLB resistance in two recombinant inbred line populations and to compare these to SLB resistance loci in other populations. The two populations used were derived from crosses between maize inbred lines H99 and B73 (HB population-142 lines) and between B73 and B52 (BB population-186 lines). They were evaluated for SLB resistance and for days from planting to anthesis (DTA) in 2005 and 2006. Two replications arranged as randomized complete blocks were assessed in each year for each population. Entry mean heritabilities for disease resistance were high for both populations (0.876 and 0.761, respectively). Quantitative trait loci (QTL) for SLB resistance were identified in bins 3.04 (two QTL), 6.01, and 8.05 in the HB population and in bin 2.07 in the BB population. No overlap of DTA and SLB resistance QTL was observed, nor was there any phenotypic correlation between the traits. A comparison of the results of all published SLB resistance QTL studies suggested that bins 3.04 and 6.01 are 'hotspots' for SLB resistance QTL.

The grapevine genome sequence suggests ancestral hexaploidization in major angiosperm phyla.

The analysis of the first plant genomes provided unexpected evidence for genome duplication events in species that had previously been considered as true diploids on the basis of their genetics. These polyploidization events may have had important consequences in plant evolution, in particular for species radiation and adaptation and for the modulation of functional capacities. Here we report a high-quality draft of the genome sequence of grapevine (Vitis vinifera) obtained from a highly homozygous genotype. The draft sequence of the grapevine genome is the fourth one produced so far for flowering plants, the second for a woody species and the first for a fruit crop (cultivated for both fruit and beverage). Grapevine was selected because of its important place in the cultural heritage of humanity beginning during the Neolithic period. Several large expansions of gene families with roles in aromatic features are observed. The grapevine genome has not undergone recent genome duplication, thus enabling the discovery of ancestral traits and features of the genetic organization of flowering plants. This analysis reveals the contribution of three ancestral genomes to the grapevine haploid content. This ancestral arrangement is common to many dicotyledonous plants but is absent from the genome of rice, which is a monocotyledon. Furthermore, we explain the chronology of previously described whole-genome duplication events in the evolution of flowering plants.

The role of barren stalk1 in the architecture of maize.

The architecture of higher plants is established through the activity of lateral meristems--small groups of stem cells formed during vegetative and reproductive development. Lateral meristems generate branches and inflorescence structures, which define the overall form of a plant, and are largely responsible for the evolution of different plant architectures. Here, we report the isolation of the barren stalk1 gene, which encodes a non-canonical basic helix-loop-helix protein required for the initiation of all aerial lateral meristems in maize. barren stalk1 represents one of the earliest genes involved in the patterning of maize inflorescences, and, together with the teosinte branched1 gene, it regulates vegetative lateral meristem development. The architecture of maize has been a major target of selection for early agriculturalists and modern farmers, because it influences harvesting, breeding strategies and mechanization. By sampling nucleotide diversity in the barren stalk1 region, we show that two haplotypes entered the maize gene pool from its wild progenitor, teosinte, and that only one was incorporated throughout modern inbreds, suggesting that barren stalk1 was selected for agronomic purposes.

Fracaso renal agudo obstructivo por masa retroperitoneal / Acute obstructive renal failure secondarty to retroperitoneal mass

El fracaso renal agudo es un proceso grave, de instauración rápida y relativamente frecuente en el medio hospitalario. Clásicamente se describen tres grupos etiológicos responsables del mismo, entre los cuales destacan las causas prerrenales. La patología obstructiva, de menor incidencia, aumenta con la edad. Se describe el caso de un paciente de 67 años que ingresa en el Servicio de Nefrología por un deterioro brusco de la función renal. Entre los síntomas iniciales, presenta hipertensión arterial (190/90) y diuresis conservada. La analítica pone de manifiesto una urea de 199 mg/dl, con creatinina de 7,7 mg/dl, sin proteinuria. La ecografía informa una hidronefrosis renoureteral bilateral con quistes simples de posible origen isquémico. Dada la ausencia de datos bioquímicos previos de insuficiencia renal, se consideran posibles causas desencadenantes del cuadro agudo. En la valoración inicial, la etiología prerrenal no tienen fundamento clínico (tensión arterial elevada, correcta función sistólica cardíaca por ecocardiografía), ni bioquímico. La ausencia de síndrome prostático y los hallazgos ecográficos tampoco justifican una obstrucción del tracto urinario. Por último, la tomografía abdomino-pélvica evidencia una masa retroperitoneal periaórtica que engloba ambos uréteres y parece desencadenante del cuadro obstructivo. Se inicia un seguimiento conjunto con el Servicio de Urología, que implanta unos catéteres «doble J» bilaterales y más adelante, interviene quirúrgicamente al paciente llevando a efecto una ureterolisis alterna de ambos uréteres. El material de biopsia informa de fibrosis retroperitoneal, y el renograma evalúa una función renal residual del 20% en riñón derecho y del 80% en el izquierdo. Ante el fracaso de las medidas quirúrgicas anteriores y como último recurso terapéutico, se ensaya al año del diagnóstico, una pauta continua con tamoxifeno (antiestrógeno) a dosis de 20 mg/dl cada 12 horas, que inicia una remisión progresiva en el tamaño de la masa, objetivada por tomografía (TAC) y resonancia magnética (RMN). La pauta se completa durante 12 meses y en este tiempo se reducen también de forma paulatina, los niveles de nitrógeno uréico y creatinina en sangre. Finalmente al término del tratamiento, la RMN evidencia la desaparición total de la fibrosis (AU)

The acute renal failure is a grave pathology, of rapid establishment and relatively frecuent in the hospital environment. We can describe three etiological group, which are responsible of himself, amongst which are emphasized the pre-renal reasons. The obstructive pathology, of minor incidence, increase with the age. It in described the case of a patient with 67 years old who is admitted in the Nephrology Service because of abrupt spoiling of the renal function. Between initials symptons, he presents arterial hipertension (190/90) and preserved diuresis. The analytics manifest: urea 199 mg/dl, creatinine 7.7 mg/dl, without proteinuria. Sonography reports a bilateral ureteral hydronephrosis with simple cyst of possible ischemia origin. In view of the absence of previus biochemicals data of renal failure, it is considered possibles reasons which start acute pattern. In initial evaluation, pre-renal etiology has not clinical basis (high blood pressure, right cardiac systole function). The absence of prostatic syndrome and sonography discovery, neither justify an urinary tract obstruction. Finally, abdominal-pelvic scan show a periaortic retroperitoneal mass which include both ureter and appear like outbreak ob obstruent patern. It is begun a combined pursuit with the Urology Service, which implant a bilateral «double J» catheter and later, operate on surgicaly the patient, carrying out an alternating ureterolysis of both ureter. The equipment of byopsy manifested a retroperitoneal fibrosis, and the renogram evaluate a residual renal function of 20% inside right kidney and 80% inside left kidney. Before the failure of the previous measuring and like last therapeutic recourse, it is tried when one year has passed from the diagnostic, a continuous guide with tamoxifen (anti-estrogen drug) in dose of 20 mg/dl each 12 hours, which begins a progressive remission in the size of the objectified mass by scan (CT) and magnetic resonance (MR). The guide is completed during 12 months and in this time, the levels of ureic nitrogen and creatinine in blood, are reduced gradually too. Finally, at the end of the treatment, the magnetic resonance demonstrate the complete disappearance of the fibrosis (AU)

OsMADS13, a novel rice MADS-box gene expressed during ovule development.

MADS-box genes have been shown to play a major role in defining plant architecture. Recently, several MADS-box genes have been reported that are highly expressed in the ovule. However, only for the Petunia genes FBP7 and FBP11 has a function in defining ovule identity been shown. We have isolated a rice MADS-box gene named OsMADS13. Expression analysis has shown that this gene is highly expressed in developing ovules. In order to facilitate a detailed characterization of rice ovule-expressed genes, a comprehensive morphological description of ovule development in rice has been performed. The predicted amino acid sequence of OsMADS13 shows significant homology with ZAG2, a maize MADS-box gene, which is also expressed mainly in the ovule. Mapping of the gene in the rice genome showed that it is located on chromosome 12, which is syntenic to two maize regions where ZAG2 and its paralogous gene ZMM1 have been mapped. Our results suggest that OsMADS13 is the ortholog of ZAG2 and ZMM1 and might play a role in rice ovule and seed development.

CaMADS1, a MADS box gene expressed in the carpel of hazelnut.

Hazelnut (Corylus avellana L.) is a species of economic interest that shows a peculiar floral biology. Unlike most of the angiosperms, which produce ovules during floral development such that they are ready for pollen at anthesis, hazelnut ovary development is delayed and triggered by compatible pollination. In order to elucidate the mechanisms regulating this unusual process and the role of the MADS box genes in ovary development, a cDNA library from pollinated styles of hazelnut was screened with a mixture of MADS box genes from different plant species. CaMADS1 (Corylus avellana MADS box), a floral-specific MADS box gene, was isolated, and characterized as belonging to the sub-family of the AGAMOUS genes. Northern blot, RT-PCR analyses and in situ hybridization experiments show a precise correlation between ovary development and CaMADS1 expression, indicating a role of this MADS box gene in the processes of floral organogenesis.

The anti-tumor effect of photodynamic therapy evaluated by bromodeoxyuridine immunohistochemistry.

The anti-tumor effect of photodynamic therapy (PDT) on mouse tumors was evaluated with bromodeoxyuridine (BrdU) immunohistochemistry. BrdU was injected into the mice intraperitoneally (40 mg/kg body weight). Immediately after injection of BrdU, PDT using a photosensitizing drug (hematoporphyrin oligomers: 20 mg/kg body weight) was carried out on the experimental group but not on the control group. BrdU labeling indices (LIs) of the tumor cells close to blood vessels and adjacent to the surrounding normal tissue were investigated. In the tumor cells close to blood vessels, the LIs of the experimental group were significantly lower than those of the control group. As for the tumor cells adjacent to the surrounding normal tissue, the LIs of the experimental group were similar to those of the control group. Thus, the effect of PDT was significant in the tumor cells close to the blood vessels, while the tumor cells adjacent to the surrounding normal tissue resisted PDT.

Characterization of transformation related genes in oral cancer cells.

A cDNA representational difference analysis (cDNA-RDA) and an arrayed filter technique were used to characterize transformation-related genes in oral cancer. From an initial comparison of normal oral epithelial cells and a human papilloma virus (HPV)-immortalized oral epithelial cell line, we obtained 384 differentially expressed gene fragments and arrayed them on a filter. Two hundred and twelve redundant clones were identified by three rounds of back hybridization. Sequence analysis of the remaining clones revealed 99 unique clones corresponding to 69 genes. The expression of these transformation related gene fragments in three nontumorigenic HPV-immortalized oral epithelial cell lines and three oral cancer cell lines were simultaneously monitored using a cDNA array hybridization. Although there was a considerable cell line-to-cell line variability in the expression of these clones, a reliable prediction of their expression could be made from the cDNA array hybridization. Our study demonstrates the utility of combining cDNA-RDA and arrayed filters in high-throughput gene expression difference analysis. The differentially expressed genes identified in this study should be informative in studying oral epithelial cell carcinogenesis.

Ritleng intubation system for treatment of congenital nasolacrimal duct obstruction.

Twenty-eight patients (34 eyes) with congenital nasolacrimal duct obstruction underwent silicone intubation with the Ritleng lacrimal intubation system. The technique involves introduction of a Prolene (Ethicon Inc, Somerville, NJ) monofilament guide thread, securely fastened to the silicone tubing, into a tubular metal probe that opens into the inferior meatus. The outcome was evaluated in terms of ease of intubation and objective success rate. Thirty-two (94%) of the 34 lacrimal systems were successfully intubated with the Ritleng system. Difficulty passing the Prolene thread through the probe and out the tip, necessitating conversion to a Crawford intubation system, was encountered in only 2 eyes (6%). The Prolene spontaneously emerged from the nose in 24 (75%) of 32 eyes, making retrieval simple and uncomplicated. The success rate for relieving signs and symptoms of obstruction was 97% (31/32) for the eyes with the Ritleng system and 100% (2/2) for the eyes with the Crawford system. Bicanalicular silicone intubation with the Ritleng intubation system is an easy and effective technique for treatment of congenital nasolacrimal duct obstruction.

MADS box genes expressed in developing inflorescences of rice and sorghum.

With the aim of elucidating the complex genetic system controlling flower morphogenesis in cereals, we have characterized two rice and two sorghum MADS box genes isolated from cDNA libraries made from developing inflorescences. The rice clones OsMADS24 and OsMADS45, which share high homology with the Arabidopsis AGL2 and AGL4 MADS box genes, are expressed in the floral meristem, in all the primordia, and in mature floral organs. High expression levels have also been found in developing kernels. The sorghum clone SbMADS1 is also homologous to AGL2 and AGL4: expression analysis and mapping data suggest that it is the ortholog of OsMADS24. The pattern of expression of SbMADS2, the other sorghum MADS box gene, suggests that it may play a role as a meristem identity gene, as does AP1 in Arabidopsis, to which it shows considerable homology. The four genes have been mapped on a rice RFLP genetic map: the results are discussed in terms of synteny among cereals.

Enhanced polymer one-step staining for proliferating cell nuclear antigen in squamous cell carcinoma of the tongue.

A rapid enhanced polymer one-step staining for proliferating cell nuclear antigen (EPOS-PCNA) in formalin fixed, paraffin embedded sections of tongue squamous cell carcinoma is described. EPOS-PCNA was compared with PCNA immunohistochemistry using the avidin-biotin complex (ABC) method, with or without autoclave pretreatment. Significant correlation of PCNA labeling index (percentage of labeled cells/total cells counted) was observed between EPOS-PCNA and the immunohistochemical protocol using autoclave pretreatment. We consider this method a reliable, timesaving technique that would be useful in quantitative histopathology and in performing double immunostaining.

Identification of genetic factors for Alachlor tolerance in maize by molecular markers analysis.

Genetic factors controlling tolerance to the herbicide Alachlor in maize were localised by means of two different strategies. In the first approach, backcross (BC) plants, derived from pollen which had been subjected to selective pressure for resistance to the herbicide, were analysed for segregation distortion at 47 RFLP loci and compared to BC plants obtained from non-selected pollen. Preferential transmission of five chromosomal regions where putative QTLs (Quantitative Trait Loci) are localised was revealed in the BC plants from selected pollen. A second approach was based on a classical linkage analysis for segregation of the same set of RFLPs and factors controlling the trait, in a BC population of 210 individuals, by means of regression analysis. This study detected seven significant loci in four genomic regions. Overall, two loci revealed both segregation distortion and association with the expression of the trait, indicating linkage to genes expressed in both gametophytic and sporophytic phase. Three chromosomal regions appeared to carry factors involved in plant tolerance to Alachlor which are not expressed in pollen. Conversely, three loci were linked to factors selectable in pollen, but did not reveal significant association with tolerance in the plant in the segregating populations.

Molecular analysis and mapping of two genes encoding maize glutathione S-transferases (GST I and GST II).

Maize glutathione S-transferase (GST) isozymes are encoded by a gene family comprising at least five genes, three of which (Gst I, II and III) have recently been isolated and sequenced. The enzymes are active as homo or heterodimers and exhibit intraspecific polymorphism including a "null" variant for the two major isoforms expressed in roots. Northern blot analyses performed on total root RNA from "null" and "plus" genotypes, using Gst I- and Gst II-specific probes, indicated that the Gst I gene controls the expression of the two major GST isoforms expressed in roots. Gst I and Gst II were mapped by RFLP analysis using an F2 population of 149 individuals previously characterized. Gst I was localized on the long arm of chromosome 8, while two putative Gst II loci were mapped to chromosome 8 (70 cM from Gst I) and 10, respectively.