High-fidelity SIM reconstruction-based super-resolution quantitative FRET imaging

Structured illumination-based super-resolution Förster resonance energy transfer microscopy(SIM-FRET)provides an approach to resolving molecular behavior localized in intricate biological structures in living cells.However,SIM reconstruction artifacts will decrease the quantitative analysis fidelity of SIMFRET signals.To address these issues,we have developed a method called HiFi spectrum optimization SIM-FRET(HiFi-SO-SIM-FRET),which uses optimized Wiener parameters in the two-step spectrum optimization to suppress sidelobe artifacts and achieve super-resolution quantitative SIM-FRET.We validated our method by demonstrating its ability to reduce reconstruction artifacts while maintaining the accuracy of FRET signals in both simulated FRET models and live-cell FRET-standard construct samples.In summary,HiFi-SO-SIM-FRET provides a promising solution for achieving high spatial resolution and reducing SIM reconstruction artifacts in quantitative FRET imaging.

Theoretical strategies for high-resolution mapping of complex genetic disorders in humans

Positional cloning of gene(s) underlying a complex disease trait poses requirement of a highresolution linkage map between the disease locus and genetic marker loci. Recent researches have shown that this may be achieved through appropriately modeling and screening linkage disequilibrium between the candidate marker locus and the major trait locus. However, these models were restricted to the circumstances where genotyping at the disease locus was feasible. The major limitations of pedigree-based linkage analyses were addressed in the light of positional cloning and positional candidate gene identification in humans. It summarizes the recent efforts in developing theories for fine-scale mapping of genes underlying complex genetic variations where the one-to-one relationship no longer exists between phenotype of the genetic disorders and the corresponding genotype.

Rate of decay in admixture linkage disequilibrium and its implication in gene mapping

Modeling linkage disequilibria (LD) between genes usually observed in admixed natural populations has been shown an effective approach in high-resolution mapping of disease genes in humans. A prerequisite to obtain accurate estimation of recombination fraction between genesat a marker locus and the disease locus using the approach is a reliable prediction of the proportion of the admixture populations. The present study suggested the use of gene frequencies to predict the estimate of the admixture proportion based on the observation that the gene frequencies are much more stable quantities than the haplotype frequencies over evolution of the population. In this paper, we advanced the theory and methods by which the decay rate of nonlinear term of LD in admixed population may be used to estimate the recombination fraction between the genes. Theoretical analysis and simulation study indicate that, the larger the difference of gene frequencies between parental populations and the more closely the

A population genetics model of linkage disequilibrium in admixed populations

Understanding linkage disequilibrium (LD) created in admixed population and the rate of decay in the disequilibrium over evolution is an important subject in population genetics theory and in disease gene mapping in human populations. The present study represents the theoretical investigation of effects of gene frequencies, levels of LD and admixture proportions of donor populations on the evolutionary dynamics of the LD of the admixed population. We examined the conditions under which the admixed population reached linkage equilibrium or the peak level of the LD. The study reveals the inappropriateness in approximating the dynamics of the LD generated by population admixture by the commonly used formula in literature. An appropriate equation for the dynamics isproposed. The distinct feature of the newly suggested formula is that the value of the nonlinear component of the LD remains constant in the first generation of the population evolution. Comparison between the predicted disequilibrium dynamics

Structured illumination-based super-resolution live-cell quantitative FRET imaging

Forster resonance energy transfer(FRET)microscopy provides unique insight into the functionality of biological systems via imaging the spatiotemporal interactions and functional state of proteins.Distinguishing FRET signals from sub-diffraction regions requires super-resolution(SR)FRET imaging,yet is challenging to achieve from living cells.Here,we present an SR FRET method named SIM-FRET that combines SR structured illumination microscopy(SIM)imaging and acceptor sensitized emission FRET imaging for live-cell quantitative SR FRET imaging.Leveraging the robust co-localization prior of donor and accepter during FRET,we devised a mask filtering approach to mitigate the impact of SIM reconstruction artifacts on quantitative FRET analysis.Compared to wide-field FRET imaging,SIM-FRET provides nearly twofold spatial resolution enhancement of FRET imaging at sub-second timescales and maintains the advantages of quantitative FRET analysis in vivo.We validate the resolution enhancement and quantitative analysis fidelity of SIM-FRET signals in both simulated FRET models and live-cell FRET-standard construct samples.Our method reveals the intricate structure of FRET signals,which are commonly distorted in conventional wide-field FRET imaging.

Des=gn of efficient simplified genomic DNA and bisulfite sequencing in large plant populations

The next generation sequencing enables generation of high resolution and high throughput data for structure sequence of any genome at a fast declining cost. This opens opportunity for population based genetic and genomic analyses. In many applications, whole genome sequencing or re-sequencing is unnecessary or prohibited by budget limits. The Reduced Representation Genome Sequencing （RRGS）, which sequences only a small proportion of the genome of interest, has been proposed to deal with the situations. Several forms of RRGS are proposed and implemented in the literature. When applied to plant or crop species, the current RRGS protocols shared a key drawback that a significantly high proportion （up to 60%） of sequence reads to be generated may be of non-genomic origin but attributed to chloroplast DNA or rRNA genes, leaving an exceptional low efficiency of the sequencing experiment. We recommended and discussed here the design of optimized simplified genomic DNA and bisulfite sequencing strategies, which may greatly improves efficiency of the sequencing experiments by bringing down the presentation of the undesirable sequencing reads to less than 10% in the whole sequence reads. The optimized RAD- seq and RRBS-seq methods are potentially useful for sequence variant screening and genotyping in large plant/crop populations.