Simplified methodology for large scale isolation of homozygous transgenic lines of lettuce

Background: Lettuce is a globally important leafy vegetable and a model plant for biotechnology due to its adaptability to tissue culture and stable genetic transformation. Lettuce is also crucial for functional genomics research in the Asteraceae which includes species of great agronomical importance. The development of transgenic events implies the production of a large number of shoots that must be differentiated between transgenic and non-transgenic through the activity of the selective agent, being kanamycin the most popular. Results: In this work we adjusted the selection conditions of transgenic seedlings to avoid any escapes, finding that threshold concentration of kanamycin was 75 mg/L. To monitor the selection system, we studied the morphological response of transgenic and non-transgenic seedlings in presence of kanamycin to look for a visual morphological marker. Several traits like shoot length, primary root length, number of leaves, fresh weight, and appearance of the aerial part and development of lateral roots were affected in non-transgenic seedlings after 30 d of culture in selective media. However, only lateral root development showed an early, qualitative and reliable association with nptII presence, as corroborated by PCR detection. Applied in successive transgenic progenies, this method of selection combined with morphological follow-up allowed selecting the homozygous presence of nptII gene in 100% of the analyzed plants from T2 to T5. Conclusions: This protocol allows a simplified scaling-up of the production of multiple homozygous transgenic progeny lines in the early generations avoiding expensive and time-consuming molecular assays

HPLC-DAD-ESI-QTOF-MS and HPLC-FLD-MS as valuable tools for the determination of phenolic and other polar compounds in the edible part and by-products of avocado

Avocado is a tropical fruit increasingly cultivated around the world due to global interest and rising consumption. Thus, there is also a surge in avocado by-products that needs assessment. The aim of this work is to compare the phenolic profile of avocado pulp, peel and seed when the fruit is at optimal ripeness for consumption and when overripe. Two analytical techniques were used: (1) HPLC-DAD-ESI-QTOF-MS was used for the first time to determine phenolic and other polar compounds in avocado peel and seed. Phenolic compounds quantified with these methods were in higher concentration in overripe than in pulp and seed of optimally ripe fruit. (2) HPLC-FLD-MS was used to specifically determine flavan-3-ols. Procyanidins to degree of polymerization 13 have been quantified singularly here for the first time. In addition, A- and B-type procyanidins from the degree of polymerization 2 to 6 were differentiated and quantified. The procyanidin concentration increased after ripening probably due to the release of tannins linked to cell-wall structures. Because of this situation and the presence of A-type procyanidins, avocado peel and seed from overripe fruit, the main by-products of avocado processing, hold interest for developing functional foods, nutraceuticals and cosmetics

Connectivity Among Populations of the Top Shell Gibbula divaricata in the Adriatic Sea

Genetic connectivity studies are essential to understand species diversity and genetic structure and to assess the role of potential factors affecting connectivity, thus enabling sound management and conservation strategies. Here, we analyzed the patterns of genetic variability in the marine snail Gibbula divaricata from five coastal locations in the central-south Adriatic Sea (central Mediterranean) and one in the adjacent northern Ionian Sea, using 21 described polymorphic microsatellite loci. Observed and expected heterozygosity varied from 0.582 to 0.635 and 0.684 to 0.780, respectively. AMOVA analyses showed that 97% of genetic variation was observed within populations. Nevertheless, significant, although small, genetic differentiation was found among nearly all of the pairwise FST comparisons. Over a general pattern of panmixia, three groups of populations were identified: eastern Adriatic populations, western Adriatic populations, and a third group represented by the single northern Ionian Sea population. Nonetheless, migration and gene flow were significant between these groups. Gibbula divaricata is thought to have a limited dispersal capacity related to its lecithotrophic trochophore larval stage. Our results indicated high levels of self-recruitment and gene flow that is mainly driven through coastline dispersion, with populations separated by the lack of suitable habitats or deep waters. This stepping-stone mode of dispersion together with the high levels of self-recruitment could lead to higher levels of population structuring and differentiation along the Adriatic Sea. Large effective population sizes and episodic events of long-distance dispersal might be responsible for the weak differentiation observed in the analyzed populations. In summary, the circulation system operating in this region creates natural barriers for dispersion that, together with life-history traits and habitat requirements, certainly affect connectivity in G. divaricata. However, this scenario of potential differentiation seems to be overridden by sporadic events of long-distance dispersal across barriers and large effective population sizes

Artificial intelligence could alert for focal skeleton/bone marrow uptake in Hodgkin’s lymphoma patients staged with FDG-PET/CT

To develop an artificial intelligence (AI)-based method for the detection of focal skeleton/bone marrow uptake (BMU) in patients with Hodgkin’s lymphoma (HL) undergoing staging with FDG-PET/CT. The results of the AI in a separate test group were compared to the interpretations of independent physicians. The skeleton and bone marrow were segmented using a convolutional neural network. The training of AI was based on 153 un-treated patients. Bone uptake significantly higher than the mean BMU was marked as abnormal, and an index, based on the total squared abnormal uptake, was computed to identify the focal uptake. Patients with an index above a predefined threshold were interpreted as having focal uptake. As the test group, 48 un-treated patients who had undergone a staging FDG-PET/CT between 2017–2018 with biopsy-proven HL were retrospectively included. Ten physicians classified the 48 cases regarding focal skeleton/BMU. The majority of the physicians agreed with the AI in 39/48 cases (81%) regarding focal skeleton/bone marrow involvement. Inter-observer agreement between the physicians was moderate, Kappa 0.51 (range 0.25–0.80). An AI-based method can be developed to highlight suspicious focal skeleton/BMU in HL patients staged with FDG-PET/CT. Inter-observer agreement regarding focal BMU is moderate among nuclear medicine physicians

Caracterizaci\uf3n del holoparasitismo de Orobanche ramosa en tomate bajo condiciones de campo

Field studies were conducted between 1999 and 2002, in Angol, Chile (37\ub045 ' S, 72\ub044 ' W) to determine the phenology of the system Orobanche ramosa \u2013 tomato ( Lycopersicon esculentum Mill.), to characterize the influence of holoparasitism on the growth and development of the host and the productive response at three planting dates (early, normal and delayed). The O. ramosa phenology includes underground and aerial phases, defining a total of six development stages: "a":: nodule; "b": nodule with initial crown roots; "c": shoot bud already visible; "d": shoot bud and crown root developed; "e": shoot development; and "f": shoot emerged from soil surface. The underground phase began with the establishment of the parasite, the first phenological event (stage "a":) being detected between 120 to 250 degree-days (DG), and the aerial phase (stage "f":) began when 550 DG of thermal requirements was surpassed. These phases occurred at flowering and fruit set of the host, respectively. The mean life cycle of the system O. ramosa - tomato was completed with 1,180 DG or 132 days after planting (DDT). Parasitism of O. ramosa altered morphological parameters and the distribution of dry matter in the host before the emergence of O. ramosa, the aerial components being most effected. It translated into important losses of production at the planting dates, which varied between 77 and 81%. Therefore, advancing or delaying the planting date does not constitute an efficient cultural practice for tomato fields to escape O. ramosa parasitism.Se realizaron estudios de campo entre 1999 y 2002, en Angol, Chile (37\ub045' lat. Sur, 72\ub044' long. Oeste) para determinar la fenolog\ueda del complejo orobanque-tomate ( Orobanche ramosa L.- Lycopersicon esculentum Mill.), caracterizar la influencia del holoparasitismo en el crecimiento y desarrollo del hospedero, y la respuesta productiva en tres \ue9pocas de plantaci\uf3n (temprana, normal y tard\ueda). La fenolog\ueda de O. ramosa comprende una fase subterr\ue1nea y otra a\ue9rea, defini\ue9ndose un total de seis estados de desarrollo: "a": n\uf3dulo; "b": n\uf3dulo con ra\uedz de escaso desarrollo; "c": aparici\uf3n de yema; "d": yema y ra\uedz desarrollada; "e": v\ue1stago desarrollado; y "f": v\ue1stago emergiendo sobre la superficie del suelo. La fase subterr\ue1nea se inici\uf3 con el establecimiento del fitopar\ue1sito, detect\ue1ndose el primer evento fenol\uf3gico (estado "a") entre 120 a 250 d\uedas grado (DG), y la fase a\ue9rea (estado "f") comenz\uf3 cuando se super\uf3 los 550 DG de requerimientos t\ue9rmicos. Dichas fases coincidieron con la floraci\uf3n y fructificaci\uf3n del hospedero, respectivamente. El ciclo de vida promedio de O. ramosa se complet\uf3 con 1.180 DG \uf3 132 d\uedas despu\ue9s del trasplante (DDT). El parasitismo alter\uf3 par\ue1metros morfol\uf3gicos y distribuci\uf3n de la materia seca en el hospedero antes de la emergencia de O. ramosa, siendo el componente a\ue9reo (follaje) el m\ue1s afectado. Ello se tradujo en importantes p\ue9rdidas de producci\uf3n en las \ue9pocas de plantaci\uf3n, las que variaron entre 77 y 81%. Por lo tanto, adelantar o retrasar la plantaci\uf3n de tomate no constituye una pr\ue1ctica cultural eficiente para escapar al efecto del parasitismo de O. ramosa

Canonical DNA minor groove insertion of bisbenzamidine-Ru(ii) complexes with chiral selectivity

We report the first Ru(ii) coordination compounds that interact with DNA through a canonical minor groove insertion mode and with selectivity for A/T rich sites. This was made possible by integrating a bis-benzamidine minor groove DNA-binding agent with a ruthenium(ii) complex. Importantly, one of the enantiomers (Δ-[Ru(bpy)2b4bpy]2+, Δ-4Ru) shows a considerably higher DNA affinity than the parent organic ligand and the other enantiomer, particularly for the AATT sequence, while the other enantiomer preferentially targets long AAATTT sites with overall lower affinity. Finally, we demonstrate that the photophysical properties of these new binders can be exploited for DNA cleavage using visible light

The role of nanoparticle structure and morphology in the dissolution kinetics and nutrient release of nitrate‑doped calcium phosphate nanofertilizers

Bio-inspired synthetic calcium phosphate (CaP) nanoparticles (NPs), mimicking the mineral component of bone and teeth, are emergent materials for sustainable applications in agriculture. These sparingly soluble salts show self-inhibiting dissolution processes in undersaturated aqueous media, the control at the molecular and nanoscale levels of which is not fully elucidated. Understanding the mechanisms of particle dissolution is highly relevant to the efcient delivery of macronutrients to the plants and crucial for developing a valuable synthesis-by-design approach. It has also implications in bone (de)mineralization processes. Herein, we shed light on the role of size, morphology and crystallinity in the dissolution behaviour of CaP NPs and on their nitrate doping for potential use as (P,N)-nanofertilizers. Spherical fully amorphous NPs and apatite-amorphous nanoplatelets (NPLs) in a core-crown arrangement are studied by combining forefront Small-Angle and Wide-Angle X-ray Total Scattering (SAXS and WAXTS) analyses. ­Ca2+ ion release rates difer for spherical NPs and NPLs demonstrating that morphology plays an active role in directing the dissolution kinetics. Amorphous NPs manifest a rapid loss of nitrates governed by surface-chemistry. NPLs show much slower release, paralleling that of ­Ca2+ ions, that supports both detectable nitrate incorporation in the apatite structure and dissolution from the core basal faces.Fondazione Cariplo 2016-0648FEDER/Ministerio de Ciencia, Innovacion y Universidades-Agencia Estatal de Investigacion (FEDER/MCIU/AEI, Spain) through the project NanoVIT RTI-2018-095794-A-C22FEDER/Ministerio de Ciencia, Innovacion y Universidades-Agencia Estatal de Investigacion (FEDER/MCIU/AEI, Spain) through the project NanoSmart RYC-2016-21042FEDER/MCIU/AEI within the Juan de la Cierva Program (JdC2017

Scientific Opinion on the safety and efficacy of Urea for ruminants

Urea supplementation to feed for ruminants provides non-protein nitrogen for microbial protein synthesis in the rumen and thus in part replaces other dietary protein sources. Urea supplementation of feed for ruminants at doses up to 1 % of complete feed DM (corresponding to 0.3 g/kg bw/day) is considered safe when given to animals with a well adapted ruminal microbiota and fed diets rich in easily digestible carbohydrates. Based on the metabolic fate of urea in ruminants, the use of urea in ruminant nutrition does not raise any concern for consumers\u2019 safety. Urea is considered to be non irritant to skin and eyes and its topical use suggests that it is not a dermal sensitiser. The risk of exposure by inhalation would be low. The substitution of protein by urea in well balanced feed for ruminants would not result in an increased environmental nitrogen load. Urea is an effective source of non-protein nitrogen substituting for dietary protein in ruminants

VALES VI: ISM enrichment in star-forming galaxies up to z∼\sim0.2 using 12^{12}CO(1-0), 13^{13}CO(1-0) and C18^{18}O(1-0) line luminosity ratios

We present Atacama Large Millimeter/sub-millimeter Array (ALMA) observations towards 27 low-redshift ($0.02< z<0.2$) star-forming galaxies taken from the Valpara\'iso ALMA/APEX Line Emission Survey (VALES). We perform stacking analyses of the $^{12}$CO($1-0$), $^{13}$CO($1-0$) and C$^{18}$O($1-0$) emission lines to explore the $L'$ ($^{12}$CO($1-0$))/$L'$($^{13}$CO($1-0$))) (hereafter $L'$($^{12}$CO)/$L'$($^{13}$CO)) and $L'$($^{13}$CO($1-0$))/$L'$(C$^{18}$O($1-0$)) (hereafter $L'$($^{13}$CO)/$L'$(C$^{18}$O) line luminosity ratio dependence as a function of different global galaxy parameters related to the star formation activity. The sample has far-IR luminosities $10^{10.1-11.9}$L$_{\odot}$ and stellar masses of $10^{9.8-10.9}$M$_{\odot}$ corresponding to typical star-forming and starburst galaxies at these redshifts. On average we find a $L'$($^{12}$CO)/$L'$($^{13}$CO) line luminosity ratio value of 16.1$\pm$2.5. Galaxies with evidences of possible merging activity tend to show higher $L'$($^{12}$CO)/$L'$($^{13}$CO) ratios by a factor of two, while variations of this order are also found in galaxy samples with higher star formation rates or star formation efficiencies. We also find an average $L'$($^{13}$CO)/$L'$(C$^{18}$O) line luminosity ratio of 2.5$\pm$0.6, which is in good agreement with those previously reported for starburst galaxies. We find that galaxy samples with high $L_{\text{IR}}$, SFR and SFE show low $L'$($^{13}$CO)/$L'$(C$^{18}$O) line luminosity ratios with high $L'$($^{12}$CO)/$L'$($^{13}$CO) line luminosity ratios, suggesting that these trends are produced by selective enrichment of massive stars in young starbursts.Comment: 16 pages, 10 figures to be published in MNRA