A native promoter–gene fusion created by CRISPR/Cas9-mediated genomic deletion offers a transgene-free method to drive oil accumulation in leaves

Achieving gain-of-function phenotypes without inserting foreign DNA is an important challenge for plant biotechnologists. Here we show that a gene can be brought under the control of a promoter from an upstream gene by deleting the intervening genomic sequence using dual-guide CRISPR/Cas9. We fuse the promoter of a non-essential photosynthesis-related gene to DIACYLGLYCEROL ACYLTRANSFERASE 2 (DGAT2) in the lipase-deficient sugar-dependent 1 mutant of Arabidopsis thaliana to drive ectopic oil accumulation in leaves. DGAT2 expression is enhanced more than twenty-fold and the triacylglycerol content increases by around thirty-fold. This deletion strategy offers a transgene-free route to engineering traits that rely on transcriptional gain-of-function, such as producing high lipid forage to increase the productivity and sustainability of ruminant farming

Effects of Dielectrophoresis on Growth, Viability and Immuno-reactivity of Listeria monocytogenes

Dielectrophoresis (DEP) has been regarded as a useful tool for manipulating biological cells prior to the detection of cells. Since DEP uses high AC electrical fields, it is important to examine whether these electrical fields in any way damage cells or affect their characteristics in subsequent analytical procedures. In this study, we investigated the effects of DEP manipulation on the characteristics of Listeria monocytogenes cells, including the immuno-reactivity to several Listeria-specific antibodies, the cell growth profile in liquid medium, and the cell viability on selective agar plates. It was found that a 1-h DEP treatment increased the cell immuno-reactivity to the commercial Listeria species-specific polyclonal antibodies (from KPL) by ~31.8% and to the C11E9 monoclonal antibodies by ~82.9%, whereas no significant changes were observed with either anti-InlB or anti-ActA antibodies. A 1-h DEP treatment did not cause any change in the growth profile of Listeria in the low conductive growth medium (LCGM); however, prolonged treatments (4 h or greater) caused significant delays in cell growth. The results of plating methods showed that a 4-h DEP treatment (5 MHz, 20 Vpp) reduced the viable cell numbers by 56.8–89.7 %. These results indicated that DEP manipulation may or may not affect the final detection signal in immuno-based detection depending on the type of antigen-antibody reaction involved. However, prolonged DEP treatment for manipulating bacterial cells could produce negative effects on the cell detection by growth-based methods. Careful selection of DEP operation conditions could avoid or minimize negative effects on subsequent cell detection performance

Diverting phenylpropanoid pathway flux from sinapine to produce industrially useful 4-vinyl derivatives of hydroxycinnamic acids in Brassicaceous oilseeds

Sinapine (sinapoylcholine) is an antinutritive phenolic compound that can account for up to 2% of seed weight in brassicaceous oilseed crops and reduces the suitability of their protein-rich seed meal for use as animal feed. Sinapine biosynthesis draws on hydroxycinnamic acid precursors produced by the phenylpropanoid pathway. The 4-vinyl derivatives of several hydroxycinnamic acids have industrial applications. For example, 4-vinyl phenol (4-hydroxystyrene) is a building block for a range of synthetic polymers applied in resins, inks, elastomers, and coatings. Here we have expressed a modified bacterial phenolic acid decarboxylase (PAD) in developing seed of Camelina sativa to redirect phenylpropanoid pathway flux from sinapine biosynthesis to the production of 4-vinyl phenols. PAD expression led to a ∼95% reduction in sinapine content in seeds of both glasshouse and field grown C. sativa and to an accumulation of 4-vinyl derivatives of hydroxycinnamic acids, primarily as glycosides. The most prevalent aglycone was 4-vinyl phenol, but 4-vinyl guaiacol, 6-hydroxy-4-vinyl guaiacol and 4-vinylsyringol (Canolol) were also detected. The molar quantity of 4-vinyl phenol glycosides was more than twice that of sinapine in wild type seeds. PAD expression was not associated with an adverse effect on seed yield, harvest index, seed morphology, storage oil content or germination in either glasshouse or field experiments. Our data show that expression of PAD in brassicaceous oilseeds can supress sinapine accumulation, diverting phenylpropanoid pathway flux into 4-vinyl phenol derivatives, thereby also providing a non-petrochemical source of this class of industrial chemicals

Self-directed growth of AlGaAs core-shell nanowires for visible light applications

Al(0.37)Ga(0.63)As nanowires (NWs) were grown in a molecular beam epitaxy system on GaAs(111)B substrates. Micro-photoluminescence measurements and energy dispersive X-ray spectroscopy indicated a core-shell structure and Al composition gradient along the NW axis, producing a potential minimum for carrier confinement. The core-shell structure formed during the growth as a consequence of the different Al and Ga adatom diffusion lengths.Comment: 20 pages, 7 figure

Effect of FYM, foliar feeding of nitrogen and deficit irrigation on drip irrigated coriander (Coriandrum sativum L.)

A field experiment was conducted to study the water use, nitrogen (N) uptake and economics of coriander (Coriandrum sativum L.) under organic enrichment, foliar feeding of N and fertigation. Drip irrigation at 80% ETc + foliar spray of 25% N (15 kg ha-1) + fertigation (45 kg ha-1) gave the highest plant height, branch plant-1, umbel plant-1, umbelets umbel-1, seed umbel-1, and test weight compared to irrigation at 80% ETc and at 60% ETc (upto flowering stage) + 80% ETc (reproductive stage) with or without foliar spray. Increased yield parameters were attributed to the highest seed yield and B:C ratio of 1.90 using 265.74 mm water. Water use efficiency (WUE) was also the highest. This treatment also recorded the highest N uptake of 52.6 kg ha-1. Further, addition of 10 t ha-1 FYM in addition to recommended nitrogen dose (60 kg ha-1) gave higher yield attributes viz., branch plant-1, umbel plant-1, seed umbel-1 and test weight as compared to recommended level of fertilizers. Enhanced yield attributes thus increased seed yield, N removal, WUE and B:C ratio. &nbsp

Effect of drip irrigation and bioregulators on yield, economics and water use of fenugreek (Trigonella foenum-graecum)

The field experiment was conducted during winter seasons of 2009–10 and 2010–11 to assess the effect of drip irrigation levels and bioregulators (thiourea and Thioglycollic acid) on fenugreek. Increasing levels of drip irrigation from 0.6 to 1.0 ETc enhanced yield attributes viz., branches plant-1, pods plant-1, pod length, seeds pod-1 and test weight. Increased yield attributes with 1.0 ETc thus, enhanced seed yield (15.53 q ha-1) and stover yield (31.54 q ha-1). However highest B:C ratio (2.11) was recorded with 0.8 ETc irrigation level and highest water use efficiency of 4.83 kg ha-1 mm with 0.6 ETc. Recommended surface irrigation treatment recorded 13.02 q ha-1 seed yield with using 540 mm of water. Whereas, fenugreek under drip irrigation at 0.6, 0.8 and 1.0 ETc used 278.16, 370.88 and 463.60 mm of water, respectively. Seed treatment of fenugreek before sowing with bio-regulator thiourea (500 ppm) for 4 h recorded higher yield attributes viz., pods plant-1, pod length and test weight, as well as, seed yield of 14.85 q ha-1 against 14.36 q ha-1 where no seed treatment was done. Thus, in turn fetched higher water use efficiency (4.00 kg ha-1 mm) and B:C ratio (2.08). Further, foliar spray with 200 ppm thioglycollic acid at vegetative and seed formation stages recorded highest seed yield of 15.01 q ha-1 over 100 ppm thioglycollic acid spray (14.59 q ha-1) or no spray treatment (14.21 q ha-1). Increased yield thus enhanced water use efficiency (3.93-4.05 kg ha-1 mm) and B:C ratio (2.06-2.11) in thioglycollic acid treated crop. &nbsp