Evaluation of gene-finding algorithms by a content-balancing accuracy index

A content-balancing accuracy index, called q(9), to evaluate gene-finding algorithms has been proposed. Here the concept of content-balancing means that the evaluation by this index is independent of the coding and non-coding composition of the sequence being evaluated. Since the coding and non-coding compositions are severely unbalanced in eukaryotic genomes, the performance of gene-finding algorithms is either over- or under-evaluated by the widely used accuracy indices, e.g., the correlation coefficient, due to the lack of content-balancing ability. Using the new accuracy index q(9), seven gene-finding algorithms, FGENES; Gene-Mark.hmm; Genie; Genescan; HMMgene; Morgan and MZEF, were compared and evaluated. It is shown that Genescan is still the best one, but with q(9)= 89%, averaged over the prediction for 195 sequences. In addition to the content-balancing ability, q(9) has the merit of having definition in all possible cases. It is also shown that the traditional specificity s(p) carries important information on the performance of the algorithm being evaluated. The set of sensitivity s(n), specificity s(p) and the accuracy q(9) constitutes a complete kit to evaluate gene-finding algorithms at nucleotide level. In addition, a graphic method to compare and evaluate gene-finding algorithms has been proposed, too. Its major advantage is that the overall performance of algorithms can be grasped quickly in a perceivable form. Additionally, the new accuracy index q(9) may be applied to evaluate the performance of weather forecast, clinical diagnosis, psychological examination and protein secondary structure prediction etc.     

Comparative sequence analysis of 634 kb of the mouse chromosome 16 region of conserved synteny with the human velocardiofacial syndrome region on chromosome 22q11.2

Mouse genomic DNA sequence extending 634 kb on proximal mouse chromosome 16 was compared to the corresponding human sequence from chromosome 22q11.2. Haploinsufficiency for this region results in velocardiofacial syndrome (VCFS) in humans. The mouse region is rearranged into three conserved blocks relative to human, but gene content and position are highly conserved within these blocks. Examination of the boundaries of one of these blocks suggested that the evolutionary chromosomal rearrangement occurred in the mouse lineage, resulting in inactivation of the mouse orthologue of ZNF74. Sequence analysis identified 21 genes and 15 ESTs. These include 2 novel genes, Srec2 and Cals2, and previously undescribed splice variants of several other genes. Exon discovery was carried out using GRAIL2, MZEF, or comparative analysis across 491 kb of conserved mouse and human sequence. Sequence comparison was highly effective, identifying every gene and nearly every exon without the high frequency of false-positive predictions seen when algorithmic methods were used alone. In combination, these procedures identified every gene with no false-positive predictions. Comparative sequence analysis also revealed regions of extensive conservation among noncoding sequences, accounting for 6% of the sequence. A library of such sequences has been established to form a resource for generalized studies of regulatory and structural elements.     

GeneMachine: gene prediction and sequence annotation

MOTIVATION: A number of free-standing programs have been developed in order to help researchers find potential coding regions and deduce gene structure for long stretches of what is essentially 'anonymous DNA'. As these programs apply inherently different criteria to the question of what is and is not a coding region, multiple algorithms should be used in the course of positional cloning and positional candidate projects to assure that all potential coding regions within a previously-identified critical region are identified. RESULTS: We have developed a gene identification tool called GeneMachine which allows users to query multiple exon and gene prediction programs in an automated fashion. BLAST searches are also performed in order to see whether a previously-characterized coding region corresponds to a region in the query sequence. A suite of Perl programs and modules are used to run MZEF, GENSCAN, GRAIL 2, FGENES, RepeatMasker, Sputnik, and BLAST. The results of these runs are then parsed and written into ASN.1 format. Output files can be opened using NCBI Sequin, in essence using Sequin as both a workbench and as a graphical viewer. The main feature of GeneMachine is that the process is fully automated; the user is only required to launch GeneMachine and then open the resulting file with Sequin. Annotations can then be made to these results prior to submission to GenBank, thereby increasing the intrinsic value of these data. AVAILABILITY: GeneMachine is freely-available for download at http://genome.nhgri.nih.gov/genemachine. A public Web interface to the GeneMachine server for academic and not-for-profit users is available at http://genemachine.nhgri.nih.gov. The Web supplement to this paper may be found at http://genome.nhgri.nih.gov/genemachine/supplement/.     

Positional characterisation of false positives from computational prediction of human splice sites

The performance of computational tools that can predict human splice sites are reviewed using a test set of EST-confirmed splice sites. The programs (namely HMMgene, NetGene2, HSPL, NNSPLICE, SpliceView and GeneID-3) differ from one another in the degree of discriminatory information used for prediction. The results indicate that, as expected, HMMgene and NetGene2 (which use global as well as local coding information and splice signals) followed by HSPL (which uses local coding information and splice signals) performed better than the other three programs (which use only splice signals). For the former three programs, one in every three false positive splice sites was predicted in the vicinity of true splice sites while only one in every 12 was expected to occur in such a region by chance. The persistence of this observation for programs (namely FEXH, GRAIL2, MZEF, GeneID-3, HMMgene and GENSCAN) that can predict all the potential exons (including optimal and sub-optimal) was assessed. In a high proportion (>50%) of the partially correct predicted exons, the incorrect exon ends were located in the vicinity of the real splice sites. Analysis of the distribution of proximal false positives indicated that the splice signals used by the algorithms are not strong enough to discriminate particularly those false predictions that occur within ± 25 nt around the real sites. It is therefore suggested that specialised statistics that can discriminate real splice sites from proximal false positives be incorporated in gene prediction programs.     

演化极端结合分支分类方法

从生物演化的逆方向考虑,提出一种聚合的分支分类运算方法,称为演化极端结合分支分类法。文章阐明其设计思路、演算步骤,并以实例具体说明其演算过程。最后以演化长度系数、合理解与合理方法等概念,对演化极端结合法进行评价。     

改进的SDS-CTAB法提取濒危植物连香树总DNA

对珍稀濒危植物连香树(Cercidiphyllum japonicum)的6种总DNA提取方法进行了对比试验,结果表明改进的SDS-CTAB法更适合于连香树总DNA提取。该方法提取的DNA经紫外消光值检测,其A260/A280为1.8532,优于CTAB法(1.4872)、SDS法(1.3552)、PVP法(1.5079)、尿素法(1.1858)和高盐低pH法(1.4534)。琼脂糖凝胶电泳和PCR扩增结果也得出同样的结论。     

用气相色谱-质谱联用比较牛耳草代谢物的提取方法

通过比较不同的提取方法对牛耳草新鲜和脱水叶片中代谢物的提取效率,旨在建立一种可以有效鉴定并分析牛耳草脱水过程中关键小分子代谢物的种类和含量变化的方法,为研究植物耐脱水分子机制提供技术方法。本研究以气相色谱-质谱联用(GC-MS)为分析方法,对复苏植物牛耳草代谢物提取方法进行比较。从提取总色谱峰数目、提取效率、代谢物保留时间和提取效率稳定性等方面比较甲醇溶液(A法)和甲醇-氯仿-水溶液(B法)两种提取方法的提取效果。对牛耳草新鲜样品提取结果表明,B法提取的总色谱峰数目多于A法;对9种共有代谢物的提取效率比较结果表明,B法的提取效率高于A法;对10种色谱峰的保留时间和提取效率的方法学考察结果表明,两者保留时间RSD(相对标准偏差)值均小于1%,A法提取效率的RSD值≤10%的比例为50%,B法的为100%。A法对干样的提取色谱峰数目远少于鲜样,而B法对干样的提取色谱峰数目和鲜样没有显著差异,保留时间RSD值均小于1%,提取效率的RSD值与鲜样没有差异,稳定性良好。     

Technique for measuring picomolar amounts of bound and unbound amino acids obtained from myogenic cell cultures of skeletal muscle

Summary A procedure is described that compares the isotope dilution method of measuring picomolar amounts of amino acids obtained from cellular extracts with a direct method of analysis. Evidence is provided that shows that the direct method is at least as accurate as the isotope dilution method. In addition the direct method is as expedient and requires but a single isotope and fewer chromatograms for analysis. A procedure also is described for selecting the appropriate conditions for dansylation and for measuring the loss of dansyl amino acid due to decomposition. This research was funded through a grant from the Muscular Dystrophy Association of Canada.     

A Concordance Method For Analyzing Categorical Time Series. An Application For The Search of Periodicities

A simple method for searching for periods in biological series is proposed. Because it is based on an auto-comparison of the observations within a series we call it the concordance method. It requires few theoritical assumptions. In fact, even the ever present stationarity condition is not used. The method is compared with competing methods based on the khi-square periodogram. It is shown that the concordance method is much better for analyzing multimodal and noisy series. Rhythms presenting simultaneously circadian and ultradian components can also be analyzed with this method.     

农杆菌质粒DNA的间接酶切鉴定

采用常规手段提酶切鉴定法,与普通大肠杆菌质粒小量抽提试剂盒提取农杆菌质粒酶切鉴定法(简称试剂盒法)和农杆菌质粒反导大肠杆菌间接酶切鉴定法(简称间接法)进行对比,发现本试验创新的试剂盒法和间接法可轻松做酶切鉴定,可为农杆菌质粒DNA提取经验不足者参考.     

电化学法快速检测微生物的发展现状及趋势

自1898年Stewart提出利用电化学法检测微生物,电化学法已发展成为一种微生物快速检测的方法。根据检测的参数不同,电化学微生物检测法可以分为阻抗微生物法和介电常数法。阻抗法主要用于食品工业中微生物的快速检测(≤10⁷ cfu/mL),尤其用于易腐食品的微生物快速检测,以期实现在其发生明显腐败之前得到检测结果。而介电常数则用于生物发酵过程中的微生物数量的快速测定,可以实现在线监测微生物数量及生物发酵过程的实时控制。电化学法由于其检测迅速、可以实现自动化检测,在工业化生产中具有广阔的应用前景。     

一种新颖的抗原信号增强方法─BT法在免疫组织化学中的应用

本文报道了一种新颖的抗原信号增强方法在免疫组织化学中的应用。该方法的特点是利用生物素标记的酪氨（ＢＴ）结合到ＨＲＰ的催化位点上来达到增强信号的目的。本研究以冰冻切片及石蜡切片中ＡＢＣ法为对照,结合微波和蛋白酶消化预处理,采用ＢＴ法检测了人扁桃体石蜡切片标本中淋巴细胞ＩｇＤ的表达。结果表明该方法检测敏感性高,当一抗稀释至１：５０００时仍能测得ＩｇＤ的表达,与对照的ＡＢＣ法相比,检测敏感性成百倍地提高。该方法能将原先只能用于冰冻切片的单抗ＩｇＤ应用于石蜡切片中。本文还对该方法的可能机制进行了探讨。本研究提示ＢＴ法在病理检验和医学研究中有良好的应用前景。     

Approximating identity-by-descent matrices using multiple haplotype configurations on pedigrees

Identity-by-descent (IBD) matrix calculation is an important step in quantitative trait loci (QTL) analysis using variance component models. To calculate IBD matrices efficiently for large pedigrees with large numbers of loci, an approximation method based on the reconstruction of haplotype configurations for the pedigrees is proposed. The method uses a subset of haplotype configurations with high likelihoods identified by a haplotyping method. The new method is compared with a Markov chain Monte Carlo (MCMC) method (Loki) in terms of QTL mapping performance on simulated pedigrees. Both methods yield almost identical results for the estimation of QTL positions and variance parameters, while the new method is much more computationally efficient than the MCMC approach for large pedigrees and large numbers of loci. The proposed method is also compared with an exact method (Merlin) in small simulated pedigrees, where both methods produce nearly identical estimates of position-specific kinship coefficients. The new method can be used for fine mapping with joint linkage disequilibrium and linkage analysis, which improves the power and accuracy of QTL mapping.     

Method for rapid screening of pesticide mineralization in soil

A method has been developed for the analysis of (14)CO(2) evolution from the mineralization of (14)C-labelled organic compounds in soil samples. The new method is less space demanding and substantially cuts down laborious manual work compared to the traditional incubation bottle method used. Furthermore, the use of scintillation cocktail is largely reduced with the new method. In the new method, (14)CO(2) is trapped in filter paper held in the lid of a 20 ml glass vial by surface tension. The trapping solution used is Ca(OH)(2), which fixates CO(2) in the filter paper and the analysis of trapped (14)CO(2) is done using the Cyclone trade mark Storage Phosphor system. The lids are placed in a 32 well holder and exposed to a phosphor screen prior to scanning in a Cyclone trade mark scanner. The new filter method has been tested and compared to results obtained using the traditional method. The results show good agreement but due to a smaller capacity for CO(2) with the filter method compared to the traditional method, the interval between sampling has to be shorter using the filter method when the CO(2) development is high. The detection limits for the filter method is higher compared to the traditional method. With the filter method, the level of radioactivity has to exceed 300 dpm before detection is possible, while the same limit for the traditional method is around 30 dpm. On the other hand, the gas trapping faster and the efficiency is higher with the filter method.     

A new colorimetric determination of D-amino acid oxidase and urate oxidase activity.

A new method of colorimetric determination of d-amino acid oxidase and urate oxidase using catalase and 4-amino-3-hydrazino-5-mercapto-1,2,4-triazole is reported. This method is based on the combination of two steps of enzyme reactions and colorimetric procedure. The values obtained by this method are satisfactorily correlated with those obtained by the dinitrophenylhydrazine method for d-amino acid oxidase activity and the ultraviolet method for urate oxidase activity and showed good reproducibility and accuracy. It is considered that the method can be useful as a method of activity determination for studying enzyme kinetics and the reaction mechanism.     

A Concordance Method For Analyzing Categorical Time Series. An Application For The Search of Periodicities

A simple method for searching for periods in biological series is proposed. Because it is based on an auto-comparison of the observations within a series we call it the concordance method. It requires few theoritical assumptions. In fact, even the ever present stationarity condition is not used. The method is compared with competing methods based on the khi-square periodogram. It is shown that the concordance method is much better for analyzing multimodal and noisy series. Rhythms presenting simultaneously circadian and ultradian components can also be analyzed with this method.     

Evaluating a new method to estimate the rate of leaf respiration in the light by analysis of combined gas exchange and chlorophyll fluorescence measurements

Day respiration (R(d)) is an important parameter in leaf ecophysiology. It is difficult to measure directly and is indirectly estimated from gas exchange (GE) measurements of the net photosynthetic rate (A), commonly using the Laisk method or the Kok method. Recently a new method was proposed to estimate R(d) indirectly from combined GE and chlorophyll fluorescence (CF) measurements across a range of low irradiances. Here this method is tested for estimating R(d) in five C(3) and one C(4) crop species. Values estimated by this new method agreed with those by the Laisk method for the C(3) species. The Laisk method, however, is only valid for C(3) species and requires measurements at very low CO(2) levels. In contrast, the new method can be applied to both C(3) and C(4) plants and at any CO(2) level. The R(d) estimates by the new method were consistently somewhat higher than those by the Kok method, because using CF data corrects for errors due to any non-linearity between A and irradiance of the used data range. Like the Kok and Laisk methods, the new method is based on the assumption that R(d) varies little with light intensity, which is still subject to debate. Theoretically, the new method, like the Kok method, works best for non-photorespiratory conditions. As CF information is required, data for the new method are usually collected using a small leaf chamber, whereas the Kok and Laisk methods use only GE data, allowing the use of a larger chamber to reduce the noise-to-signal ratio of GE measurements.     

Utilization of mixtures of carbon compounds by yeasts

As a tool for the classification of yeasts a method is presented for determining the aerobic utilization of each of 28 carbon compounds distributed over 4 mixtures containing respectively 10, 7, 7 and 4 of these compounds. Paper chromatography is used to follow the eventual disappearance of each individual carbon compound from the medium. This method is compared to the well-known standard method where a growth response to singly offered compounds in liquid medium is used for biochemical characterization. The proposed method has two advantages: results are obtained more rapidly and with less work, and the paper-chromatographic method may reveal a time sequence of utilization. Both the new method and the established standard method were applied to eight yeasts. Although the backgrounds are different, the experimental results were similar. The method presented is suitable for the differentiation of species as well as of strains within one species. It should be considered for use in taxonomy.     

Interval Estimation for Ratios in Bioequivalence Trials

The problem for assessment of equivalence in variability of bioavailability between two drug products is considered. An exact confidence region for the ratio between intrasubject variabilities is derived when the intersubject variance is known. When the intersubject variance is unknown, a large sample approximation is considered. The proposed method for assessing equivalence in variability of bioavailability appears to be asymptotically uncorrelated with the sample mean ratio for average bioavailabilty. As a result, the proposed method in conjunction with the sample mean ratio method can be utilized for assessing population bioequivalence. An example concerning a bioequivalence trial with 24 healthy volunteers is presented to illustrate the proposed method.     

A Confidence Region Approach for Assessing Equivalence in Variability of Bioavailability

The problem for assessment of equivalence in variability of bioavailability between two drug products is considered. Similar to the method for assessing bioequivalence in average bioavailability proposed by Chow and Shao (1990), an exact confidence region approach is derived when the intersubject variance is known. When the intersubject variance is unknown, a large sample approximation is considered. The proposed method for assessing equivalence of variability of bioavailability appears to be asymptotically uncorrelated with that of Chow and Shao (1990) for average bioavailability. As a result, the proposed method in conjunction with the method proposed by Chow and Shao (1990) constitutes a confidence region approach for assessing population bioequivalence. An example concerning a bioequivalence trial with 24 healthy volunteers is provided to illustrate the proposed method.