香樟转录组基因密码子偏好性分析

为了解香樟基因密码子偏好性,该文以NCBI网站中香樟转录组数据为材料,利用生物信息学手段评价转录组数据质量,选取高质量数据的转录组,去除低质量序列,组装转录组,预测基因结构,再利用自编perl脚本提取以AUG开头的基因序列37 Mb序列34 931个基因,进一步利用CodonW分析基因密码子偏好性。结果表明:GC含量的变化范围为0.273～0.742,均值为0.452; ENC的范围为26.29～61.00,均值为52.76; CAI的范围为0.064～0.401,均值为0.199; RSCU值大于1的密码子数目为27个,其中以U或A结尾的有22个; 中性分析表明,小部分基因在对角线上,大多数基因偏离对角线; ENC-plot分析表明小部分基因在标准曲线上,大多数基因偏离标准曲线。上述研究结果表明,香樟基因的密码子偏好性比较弱,密码子常以A/U结尾; 突变和选择两者都在密码子偏好中起作用,而选择作用更大; 最终确定了GUU、CAG、GAA、UCU、GCU、GGU为最优密码子,通过对目标基因密码子的校正,提高表达效率,从而为利用基因工程技术改良香樟重要性状奠定了基础。     

Selection for minimization of translational frameshifting errors as a factor in the evolution of codon usage

In a wide range of genomes, it was observed that the usage of synonymous codons is biased toward specific codons and codon patterns. Factors that are implicated in the selection for codon usage include facilitation of fast and accurate translation. There are two types of translational errors: missense errors and processivity errors. There is considerable evidence in support of the hypothesis that codon usage is optimized to minimize missense errors. In contrast, little is known about the relationship between codon usage and frameshifting errors, an important form of processivity errors, which appear to occur at frequencies comparable to the frequencies of missense errors. Based on the recently proposed pause-and-slip model of frameshifting, we developed Frameshifting Robustness Score (FRS). We used this measure to test if the pattern of codon usage indicates optimization against frameshifting errors. We found that the FRS values of protein-coding sequences from four analyzed genomes (the bacteria Bacillus subtilis and Escherichia coli, and the yeasts Saccharomyces cerevisiae and Schizosaccharomyce pombe) were typically higher than expected by chance. Other properties of FRS patterns observed in B. subtilis, S. cerevisiae and S. pombe, such as the tendency of FRS to increase from the 5′- to 3′-end of protein-coding sequences, were also consistent with the hypothesis of optimization against frameshifting errors in translation. For E. coli, the results of different tests were less consistent, suggestive of a much weaker optimization, if any. Collectively, the results fit the concept of selection against mistranslation-induced protein misfolding being one of the factors shaping the evolution of both coding and non-coding sequences.     

Analysis of the Codon Usage Pattern in the Vibrio Cholerae Genome

Abstract

The codon usage in the Vibrio cholerae genome is analyzed in this paper. Although there are much more genes on the chromosome 1 than on chromosome 2, the codon usage patterns of genes on the two chromosomes are quite similar, indicating that the two chromosomes may have coexisted in the same cell for a very long history. Unlike the base frequency pattern observed in other genomes, the G+C content at the third codon position of the V. cholerae genome varies in a rather small interval. The most notable feature of codon usage of V. cholerae genome is that there is a fraction of genes show significant bias in base choice at the second codon position. The 2006 known genes can be classified into two clusters according to the base frequencies at this position. The smaller cluster contains 227 genes, most of which code for proteins involved in transport and binding functions. The encoding products of these genes have significant bias in amino acids composition as compared with other genes. The codon usage patterns for the 1836 function unknown ORFs are also analyzed, which is useful to study their functions.     

Horizontal gene transfer contributes to the wide distribution and evolution of type II restriction-modification systems

Restriction modification (RM) systems serve to protect bacteria against bacteriophages. They comprise a restriction endonuclease activity that specifically cleaves DNA and a corresponding methyltransferase activity that specifically methylates the DNA, thereby protecting it from cleavage. Such systems are very common in bacteria. To find out whether the widespread distribution of RM systems is due to horizontal gene transfer, we have compared the codon usages of 29 type II RM systems with the average codon usage of their respective bacterial hosts. Pronounced deviations in codon usage were found in six cases:EcoRI,EcoRV,KpnI,SinI,SmaI, andTthHB81. They are interpreted as evidence for horizontal gene transfer in these cases. As the methodology is expected to detect only one-fourth to one-third of all horizontal gene transfer events, this result implies that horizontal gene transfer had a considerable influence on the distribution and evolution of RM systems. In all of these six cases the codon usage deviations of the restriction enzyme genes are much more pronounced than those of the methyltransferase genes. This result suggests that in these cases horizontal gene transfer had occurred sequentially with the gene for the methyltransferase being first acquired by the cell. This can be explained by the fact that an active restriction endonuclease is highly toxic in cells whose DNA is not protected from cleavage by a corresponding methyltransferase.     

Allelic frequencies of an ovine scrapie susceptibility gene

Abstract

The allelic frequencies of an ovine gene associated with susceptibility to scrapie was analyzed in a sample of 30 scrapie affected sheep and 545 clinically normal sheep from 12 flocks. The allele encoding glutamine at codon 171 occurred at a frequency of 0.76 in the overall population. All 30 scrapie affected sheep were homozygous for glutamine at codon 171. This genotype was observed in 56.5% of the clinically normal sheep. None of the 30 scrapie affected sheep carried the allele encoding Valine at codon 136 although this allele was observed in 2/12 flocks sampled.     

DNA mismatch repair and synonymous codon evolution in mammals

It has been suggested that the differences in synonymous codon use between mammalian genes within a genome are due to differences in the efficiency of DNA mismatch repair. This hypothesis was tested by developing a model of mismatch repair, which was used to predict the expected relationship between the rate of substitution and G+C content at silent sites. It was found that the silent-substitution rate should decline with increasing G+C content over most of the G+C-content range, if it is assumed that mismatch repair is G+C biased, an assumption which is supported by data. This prediction was then tested on a set of 58 primate and artiodactyl genes. There was no evidence of a direct decline in substitution rate with increasing G+C content, for either twofold- or fourfold-degenerate sites. It was therefore concluded that variation in the efficiency of mismatch repair is not responsible for the differences in synonymous codon use between mammalian genes. In support of this conclusion, analysis of the model also showed that the parameter range over which mismatch repair can explain the differences in synonymous codon use between genes is very small.      

Theoretical Analysis of Computer Simulations of Sorption in a Cylindrical Micropore

Abstract

The simulation of krypton in a cylindrical pore with atomically rough walls is reconsidered. Distributions of gas-gas, gas-solid and total energy are presented and discussed in terms of their ability to characterize the adsorbed phase, especially by assigning sorbed atoms to layers within the pore. The calculation of the chemical potential of the sorbed phase from the distributions of the total energy per particle is presented and an approximate method of splitting the chemical potential into contributions due to the gas-gas and gas-solid energies is suggested and tested against the simulation data. It is found that the approximation works reasonably well for coverages up to monolayer, but shows significant deviations from the simulated values at coverages corresponding to the nearly full pore.     

Degrees of Divergence in the E. coli Genome From Correlations Between Dinucleotide,Trinucleotide and Codon Frequencies

Abstract

Oligonucleotide and codon frequencies have been determined in published sequences of E. coli DNA totaling 103,100bp with 18,459 reading frame trinucleotides; corresponding to 2.5% of the total genome. Dinucleotide frequencies are in excellent agreement with those determined by nearest neighbor chemical analysis, indicating the computer count of a limited sampling to be a good representation of the overall frequencies in total genomic DNA. The distinctive nonrandom codon pattern is found to be uniformly distributed and contributes to a distinctive nonrandom oligonucleotide pattern; enabling correlations between frequency levels to be extended beyond reading frame sequences. Correlation analysis indicates a surprisingly high degree of correlation everywhere in the genome. Coefficients of correlation between oligonucleotide frequencies overall and those in specific segments vary as follows: primary strands of individual coding sequences >0.9> lambda DNA> noncoding, non-RNA>φiX174 DNA> complementary strands> RNA genes ?0.6> transposon-insertion elements> T₇DNA? eukaryotic sequences ?0. It is concluded that this high degree of oligonucleotide and codon correspondence in E. coli reflects the widespread distribution of remnants of an early and slowly changing codon pattern that has been continually dispersed by duplication-divergence processes, leading to the present genome.     

Doublet frequencies and codon weighting in the DNA ofEscherichia coli and its phages

Summary A compilation of nucleic acid sequences fromE.coli and its phages has been analysed for the frequency of occurrence of nearest neighbour base doublets and codons. Several statistically significant deviations from random are found in both doublet and codon frequencies. The deviations inE.coli also appear to occur in and in the coat protein gene of MS2, whereas T4 and other parts of the MS2 genome show different sequence properties. These and other findings are discussed in relation to the hypothesis that rapidity of translation of mRNAs in theE. coli system is dependent on doublet frequency and codon usage patterns.     

Isolation and characterization of microsatellite loci from the endangered Mediterranean sponge Spongia agaricina (Demospongiae: Dictyoceratida)

The abundance of the bath sponge Spongia agaricina has decreased drastically in recent years and it is now considered an endangered species under Annex 3 of Bern and Barcelona conventions. We describe eight microsatellite markers and present data on their allelic variation and utility as high resolution genetic markers. We analyzed 36 individuals from two populations and found that the number of alleles per locus ranged between 1 and 7. Observed heterozygosity ranged from 0 to 0.72. We found deviations from Hardy–Weinberg expectations for some loci. We exclusively detected null alleles for those loci that deviated from Hardy–Weinberg expectations. Also, distributions of allele frequencies differed significantly between the two populations, making them suitable for population genetic analyses.     

The Codon-Degeneracy Model of Molecular Evolution

Mitochondrial genetic codons can be categorized by four patterns of nucleotide-site degeneracy based on varying combinations of twofold- or nondegenerate sites at first codon positions and twofold- or fourfold-degenerate sites at third codon positions. Herein, a model of molecular evolution is introduced that uses these patterns to calculate expected substitution frequencies for each codon position and substitution type relative to overall number of synonymous or nonsynonymous substitutions. Regions of the pocket gopher cytochrome oxidase subunit I (COI) and cytochrome b (cyt-b) genes are analyzed using this model. Chi-square distributions are used to produce relative goodness-of-fit (GF) scores for measuring the difference between substitution frequencies predicted by the codon-degeneracy model (CDM), and frequencies inferred using a well-supported phylogenetic tree of closely related species. The GF scores for expected and observed synonymous (GF_syn= 0.429, p= 0.807) and nonsynonymous (GF_ns= 2.309, p= 0.679) substitution frequencies resulted in a failure to reject the CDM as a null hypothesis for the molecular evolution of COI and cyt-b in pocket gophers. Alternative tree topologies and calculations of transition bias for these data result in higher GF scores. Received: 25 March 1999 / Accepted: 17 September 1999     

Monte Carlo dose in a prosthesis phantom based on exact geometry vs streak artefact contaminated CT data as benchmarked against Gafchromic film measurements

PurposeIn radiotherapy, accurate calculation of patient radiation dose is very important for good clinical outcome. In the presence of metallic implants, the dose calculation accuracy could be compromised by metal artefacts generated in computed tomography (CT) images of patients. This study investigates the influence of metal-induced CT artefacts on MC dose calculations in a pelvic prosthesis phantom.MethodsA pelvic phantom containing unilateral Ti prosthesis was CT-scanned and accurate Hounsfield unit (HU) values were assigned to known materials of the phantom as opposed to HU values produced through the artefact CT images of the phantom. Using the DOSXYZnrc MC code, dose calculations were computed in the phantom model constructed from the original CT images containing the artefacts and artefact-free images made from the exact geometry of the phantom with known materials. The dose calculations were benchmarked against Gafchromic EBT3 film measurements using 15 MeV electron and 10 MV photon beams.ResultsThe average deviations between film and MC dose data decreased from 3 ± 2% to 1 ± 1% and from about 6 ± 2% to 3 ± 1% for the artefact and artefact-free phantom models against film data for the electron and photon fields, respectively.ConclusionsFor the Ti prosthesis phantom, the presence of metal-induced CT artefacts could cause dose inaccuracies of about 3%. Construction of an artefact-free phantom model made from the exact geometry of the phantom with known materials to overcome the effect of artefacts is advantageous compared to using CT data directly of which the exact tissue composition is not well-known.     

Amino Acid Usage Is Asymmetrically Biased in AT- and GC-Rich Microbial Genomes

Introduction

Genomic base composition ranges from less than 25% AT to more than 85% AT in prokaryotes. Since only a small fraction of prokaryotic genomes is not protein coding even a minor change in genomic base composition will induce profound protein changes. We examined how amino acid and codon frequencies were distributed in over 2000 microbial genomes and how these distributions were affected by base compositional changes. In addition, we wanted to know how genome-wide amino acid usage was biased in the different genomes and how changes to base composition and mutations affected this bias. To carry this out, we used a Generalized Additive Mixed-effects Model (GAMM) to explore non-linear associations and strong data dependences in closely related microbes; principal component analysis (PCA) was used to examine genomic amino acid- and codon frequencies, while the concept of relative entropy was used to analyze genomic mutation rates.

Results

We found that genomic amino acid frequencies carried a stronger phylogenetic signal than codon frequencies, but that this signal was weak compared to that of genomic %AT. Further, in contrast to codon usage bias (CUB), amino acid usage bias (AAUB) was differently distributed in AT- and GC-rich genomes in the sense that AT-rich genomes did not prefer specific amino acids over others to the same extent as GC-rich genomes. AAUB was also associated with relative entropy; genomes with low AAUB contained more random mutations as a consequence of relaxed purifying selection than genomes with higher AAUB.

Conclusion

Genomic base composition has a substantial effect on both amino acid- and codon frequencies in bacterial genomes. While phylogeny influenced amino acid usage more in GC-rich genomes, AT-content was driving amino acid usage in AT-rich genomes. We found the GAMM model to be an excellent tool to analyze the genomic data used in this study.     

Inferring Adaptive Codon Preference to Understand Sources of Selection Shaping Codon Usage Bias

Alternative synonymous codons are often used at unequal frequencies. Classically, studies of such codon usage bias (CUB) attempted to separate the impact of neutral from selective forces by assuming that deviations from a predicted neutral equilibrium capture selection. However, GC-biased gene conversion (gBGC) can also cause deviation from a neutral null. Alternatively, selection has been inferred from CUB in highly expressed genes, but the accuracy of this approach has not been extensively tested, and gBGC can interfere with such extrapolations (e.g., if expression and gene conversion rates covary). It is therefore critical to examine deviations from a mutational null in a species with no gBGC. To achieve this goal, we implement such an analysis in the highly AT rich genome of Dictyostelium discoideum, where we find no evidence of gBGC. We infer neutral CUB under mutational equilibrium to quantify “adaptive codon preference,” a nontautologous genome wide quantitative measure of the relative selection strength driving CUB. We observe signatures of purifying selection consistent with selection favoring adaptive codon preference. Preferred codons are not GC rich, underscoring the independence from gBGC. Expression-associated “preference” largely matches adaptive codon preference but does not wholly capture the influence of selection shaping patterns across all genes, suggesting selective constraints associated specifically with high expression. We observe patterns consistent with effects on mRNA translation and stability shaping adaptive codon preference. Thus, our approach to quantifying adaptive codon preference provides a framework for inferring the sources of selection that shape CUB across different contexts within the genome.     

Pyrrolo[3,2-c]pyridine derivatives with potential inhibitory effect against FMS kinase: in vitro biological studies

A series of eighteen pyrrolo[3,2-c]pyridine derivatives were tested for inhibitory effect against FMS kinase. Compounds 1e and 1r were the most potent among all the other tested analogues (IC₅₀?=?60?nM and 30?nM, respectively). They were 1.6 and 3.2 times, respectively, more potent than our lead compound, KIST101029 (IC₅₀?=?96?nM). Compound 1r was tested over a panel of 40 kinases including FMS, and exerted selectivity against FMS kinase. It was further tested against bone marrow-derived macrophages (BMDM) and its IC₅₀ was 84?nM (2.32-fold more potent than KIST101029 (IC₅₀?=?195?nM)). Compound 1r was also tested for antiproliferative activity against a panel of six ovarian, two prostate, and five breast cancer cell lines, and its IC₅₀ values ranged from 0.15–1.78?µM. It possesses also the merit of selectivity towards cancer cells than normal fibroblasts.     

Mathematical studies of the interaction of respiratory gases with whole blood II. CO2 absorption

Some progress has been made on the problem of the interaction of respiratory gases with whole blood. A working mathematical model for the O₂−CO₂ interaction phenomena has been developed from mathematical studies of the data. The Edsall-Wyman (1958) model for CO₂ absorption is improved upon in this paper by consolidating it with the O₂ absorption model developed in paper I of this set (Bernard, S. R.,Bull. Math. Biophysics,22, 391–415, 1960). This improved model assumed the effect of O₂ on CO₂ absorption is mediated through the electrical charge possessed by the hemoglobin molecule,i.e., O₂ molecules bound to hemoglobin displace protons from the hemoglobin thereby increasing the negative charge on the hemoglobin and at the same time increasing the acidity of the solution. The model is tested against the data.     

Inline index helped in cleaning up data contamination generated during library preparation and the subsequent steps

Background

High-throughput sequencing involves library preparation and amplification steps, which may induce contamination across samples or between samples and the environment.

Methods

We tested the effect of applying an inline-index strategy, in which DNA indices of 6 bp were added to both ends of the inserts at the ligation step of library prep for resolving the data contamination problem.

Results

Our results showed that the contamination ranged from 0.29 to 1.25% in one experiment and from 0.83 to 27.01% in the other. We also found that contamination could be environmental or from reagents besides cross-contamination between samples.

Conclusions

Inline-index method is a useful experimental design to clean up the data and address the contamination problem which has been plaguing high-throughput sequencing data in many applications.

     

Same-strand overlapping genes in bacteria: compositional determinants of phase bias

Background

Same-strand overlapping genes may occur in frameshifts of one (phase 1) or two nucleotides (phase 2). In previous studies of bacterial genomes, long phase-1 overlaps were found to be more numerous than long phase-2 overlaps. This bias was explained by either genomic location or an unspecified selection advantage. Models that focused on the ability of the two genes to evolve independently did not predict this phase bias. Here, we propose that a purely compositional model explains the phase bias in a more parsimonious manner. Same-strand overlapping genes may arise through either a mutation at the termination codon of the upstream gene or a mutation at the initiation codon of the downstream gene. We hypothesized that given these two scenarios, the frequencies of initiation and termination codons in the two phases may determine the number for overlapping genes.

Results

We examined the frequencies of initiation- and termination-codons in the two phases, and found that termination codons do not significantly differ between the two phases, whereas initiation codons are more abundant in phase 1. We found that the primary factors explaining the phase inequality are the frequencies of amino acids whose codons may combine to form start codons in the two phases. We show that the frequencies of start codons in each of the two phases, and, hence, the potential for the creation of overlapping genes, are determined by a universal amino-acid frequency and species-specific codon usage, leading to a correlation between long phase-1 overlaps and genomic GC content.

Conclusion

Our model explains the phase bias in same-strand overlapping genes by compositional factors without invoking selection. Therefore, it can be used as a null model of neutral evolution to test selection hypotheses concerning the evolution of overlapping genes.

Reviewers

This article was reviewed by Bill Martin, Itai Yanai, and Mikhail Gelfand.