CRISPR-Cas12a assisted precise genome editing of Mycolicibacterium neoaurum.

Efficient and convenient genetic manipulation of mycobacteria, important microorganisms in human healthcare and the pharmaceutical industry, is limited. In this study, using a model strain Mycolicibacterium neoaurum ATCC 25795, the classical bacterium for the production of valuable steroidal pharmaceuticals, a genome editing system employing CRISPR-Cas12a to achieve efficient and precise genetic manipulation has been developed. Targeted genome mutations could be easily achieved by the CRISPR-Cas12a system without exogenous donor templates, assisted by innate non-homologous end-joining (NHEJ).

Immediate, multiplexed and sequential genome engineering facilitated by CRISPR/Cas9 in Saccharomyces cerevisiae.

A method called Cas-3P allowing for immediate, multiplexed and sequential genome engineering was developed using one plasmid expressing Cas9 and three marked plasmid backbones (P1, P2 and P3) for guide RNA (gRNA) expression. The three marked gRNA plasmid backbones were recurred in a P1-P2-P3 order for sequential gene targeting, without construction of any additional plasmid and elimination of gRNA plasmid by induction in each round. The efficiency of direct gRNA plasmid curing mediated by Cas-3P was more than 40% in sequential gene targeting.

Significantly Enhanced Production of Patchoulol in Metabolically Engineered .

Patchoulol, a natural sesquiterpene compound, is widely used in perfumes and cosmetics. Several strategies were adopted to enhance patchoulol production in : (i) farnesyl pyrophosphate (FPP) synthase and patchoulol synthase were fused to increase the utilization of FPP precursor; (ii) expression of the limiting genes of the mevalonate pathway was enhanced; (iii) squalene synthase was weakened by a glucose-inducible promoter of (promoter for hexose transporter) to reduce metabolic flux from FPP to ergosterol; and (iv) farnesol biosynthesis was inhibited to decrease the consumption of FPP. Glucose was used to balance the trade-off between the competitive squalene and patchoulol pathways.

CRISPR/Cpf1 facilitated large fragment deletion in Saccharomyces cerevisiae.

In this study, we focused on the applicability of CRISPR/Cpf1 in genome simplification of Saccharomyces cerevisiae and established a CRISPR/Cpf1 assisted method for rapid markerless large fragment deletion to facilitate laboratory evolution of geome of S. cerevisiae by rational genetic engineering. This method uses a Cpf1 expression plasmid and a crRNA array expression plasmid.

Cpf1-assisted efficient genomic integration of in vivo assembled DNA parts in Saccharomyces cerevisiae.

Objectives: To test the applicability of Cpf1 from Francisella novivida in genomic integration of in vivo assembled DNA parts in Saccharomyces cerevisiae.

Results: An easy-to-use vector toolkit, containing a CEN6/ARS4 plasmid expressing Cpf1 from Francisella novivida (FnCpf1) and a 2 μ plasmid for crRNA or crRNA array expressing, was constructed for Cpf1-assisted genomic integration in S. cerevisiae.

Epigenetic silencing of tumor suppressor long non-coding RNA BM742401 in chronic lymphocytic leukemia.

BM742401 is a tumor suppressor lncRNA downregulated in gastric cancer. As the promoter region and the entire transcript are embedded in a CpG island, we postulated that BM742401 is a tumor suppressor lncRNA inactivated by DNA methylation in chronic lymphocytic leukemia (CLL). The promoter of BM742401 was unmethylated in normal controls including three each of normal bone marrow, peripheral blood buffy coats, and CD19-sorted peripheral B-cells, but methylated in four (57.

[Estimating canopy water content in wheat based on new vegetation water index].

Moisture content is an important indicator for crop water stress condition, timely and effective monitoring crop water content is of great significance for evaluate crop water deficit balance and guide agriculture irrigation. In order to improve the saturated problems of different forms of typical NDWI (Normalized Different Water Index), we tried to introduce EVI (Enhanced Vegetation Index) to build new vegetation water indices (NDWI#) to estimate crop water content. Firstly, PROSAIL model was used to study the saturation sensitivity of NDWI, and NDWI# to canopy water content and LAI (Leaf Area Index).

[The new method monitoring crop water content based on NIR-Red spectrum feature space].

Moisture content is an important index of crop water stress condition, timely and effective monitoring of crop water content is of great significance for evaluating crop water deficit balance and guiding agriculture irrigation. The present paper was trying to build a new crop water index for winter wheat vegetation water content based on NIR-Red spectral space. Firstly, canopy spectrums of winter wheat with narrow-band were resampled according to relative spectral response function of HJ-CCD and ZY-3.

Assessment of the AquaCrop model for use in simulation of irrigated winter wheat canopy cover, biomass, and grain yield in the North China Plain.

Improving winter wheat water use efficiency in the North China Plain (NCP), China is essential in light of current irrigation water shortages. In this study, the AquaCrop model was used to calibrate, and validate winter wheat crop performance under various planting dates and irrigation application rates. All experiments were conducted at the Xiaotangshan experimental site in Beijing, China, during seasons of 2008/2009, 2009/2010, 2010/2011 and 2011/2012.

[Estimation of winter wheat protein content based on new indexes].

Wheat protein content is an important indicator often employed in wheat sale price. Spectral indexes and concurrent winter wheat protein content (WWPC) samples were obtained across three years. Data from 2008/2009 and 2009/2010 were utilized to build the new ratio indexes and product indexes, and then selected grey relational method and partial least squares method were used to improve the estimation accuracy of WWPC, data from 2011/2012 was utilized to validate model.

Flow-injection determination of hydrogen peroxide based on fluorescence quenching of chromotropic acid catalyzed with Fe(II).

Flow-injection analysis system (FIA system), which was based on Fe(II)-catalyzed oxidation of chromotropic acid with hydrogen peroxide, was developed for the determination of hydrogen peroxide. The chromotropic acid has a fluorescence measured at lambda(em)=440 nm (emission wavelength) with lambda(ex)=235 nm (excitation wavelength), and the fluorescence intensity at lambda(em)=440 nm quietly decreased in the presence of hydrogen peroxide and Fe(II), which was caused by Fe(II)-catalyzed oxidation of chromotropic acid with hydrogen peroxide. By measuring the difference of fluorescence intensity, hydrogen peroxide (1.

Trace and ultratrace analysis methods for the determination of phosphorus by flow-injection techniques.

Trace (< or =1 mg/l or 30 microM) and ultratrace (< or =1 microg/l or 30 nM) analysis methods for phosphorus determination by flow-injection analysis are reviewed. Most of the methods cited in this review are fundamentally based on the reaction of orthophosphate with molybdate to form heteropoly acids, such as molybdenum yellow and molybdenum blue, and some of the methods are based on the formation of such secondary reactions as ion associates and their aggregates with bulky cations, such as cationic dyes and quaternary ammonium ions. The heteropoly acids themselves can be measured by spectrophotometry, and the ion associate formed with a cationic dye, Malachite Green (MG), can be measured based on the coloration of MG.

[Analysis of part of M gene and genotyping for M segment of hantavirus detected from HFRS patients' sera in Qingdao region during 2000 - 2003].

Objective: To study the molecular epidemiological characteristics of hantavirus seen during 2000-2003 in Qingdao region of Shandong province.

Methods: Sera were collected from 64 patients with hemorrhagic fever with renal syndrome (HFRS) and viral RNA was extracted from the sera. HTN and SEO universal primers were designed as outer primers and HTN and SEO specific primers as inner primers.