An interactive microcomputer program for the computation of most probable number

An interactive computer program for the rapid computation of Most Probable Numbers (MPN) with 95% confidence intervals and goodness-of-fit, is presented. The program, written in ‘MICROSOFT BASIC’ employs an iterative algorithm based on a modification of the Newton-Raphson method and accomodates any number of replicates up to ten dilutions. It is applicable to most microcomputers with little or no modification. Since the computed MPN, confidence interval,a nd goodness-of-fit tests are displayed simultaneously, MPN tables are no longer required.     

DNABIND: an interactive microcomputer program searching for nucleotide sequences that may code for conserved DNA-binding protein motifs

This paper presents a simple program for interactive searchingfor nucleotide sequences that may code for the helix—turn—helix,zinc finger or leucine zipper motifs in proteins. The helix—turn—helixmotifs are predicted using the recently published method ofDodd and Egan, while zinc fingers and leucine zippers are searchedfor by our original methods. DNABIND is shown to detect allfour known helix—turn—helix motifs in bacteriophagelambda genes and both zinc fingers of the adrl gene of yeast.     

Dynamics of mismatched base pairs in DNA

The structural dynamics of mismatched base pairs in duplex DNA have been studied by time-resolved fluorescence anisotropy decay measurements on a series of duplex oligodeoxynucleotides of the general type d[CGG(AP)GGC].d[GCCXCCG], where AP is the fluorescent adenine analogue 2-aminopurine and X = T, A, G, or C. The anisotropy decay is caused by internal rotations of AP within the duplex, which occur on the picosecond time scale, and by overall rotational diffusion of the duplex. The correlation time and angular range of internal rotation of AP vary among the series of AP.X mismatches, showing that the native DNA bases differ in their ability to influence the motion of AP. These differences are correlated with the strength of base-pairing interactions in the various AP.X mismatches. The interactions are strongest with X = T or C. The ability to discern differences in the strength of base-pairing interactions at a specific site in DNA by observing their effect on the dynamics of base motion is a novel aspect of the present study. The extent of AP stacking within the duplex is also determined in this study since it influences the excited-state quenching of AP. AP is thus shown to be extrahelical in the AP.G mismatch. The association state of the AP-containing oligodeoxynucleotide strand is determined from the temperature-dependent tumbling correlation time. An oligodeoxynucleotide triplex is formed with a particular base sequence in a pH-dependent manner.     

Excited states of the DNA base pairs

The results of calculations of the first π-electronic states of the DNA base pairs with the SCF-MO-LCAO method both without taking into account configuration interaction and taking into account all the singly excited configurations are presented. The first excited singlet state and the first triplet state of both pairs are shown to be, independently of an approximation, the states where the excitation is localized on one of the bases.     

SEQ-ED: an interactive computer program for editing, analysis and storage of long DNA sequences

The rapidly growing body of sequenced DNA demands efficientcomputer programs for its analysis and storage. The programdescribed in this paper, SEQ-ED, has been designed to handlea large number of DNA sequences up to 200 kilobases [kb] longstored in a sequence library. In order to minimize the requiredstorage space, the sequences are stored in a compressed formatusing three binary digits per base. In the development of thisprogram, special care has been given to make it easy to usefor molecular biologists without any previous computer experience. Received on September 10, 1984; accepted on October 30, 1984     

Searching for tRNA genes in DNA sequences--an IBM microcomputer program

A user-friendly program that enables fast and exhaustive searchingfor tRNA genes is presented. The program can run on IBM personalmicrocomputers and comprises modules for sequence editing andGenBank database access. Received on June 13, 1989; accepted on August 30, 1989     

PEGASE: a machine language program for DNA sequence analysis on Apple II microcomputer using a binary coding of nucleotides

The core of a 6502 machine language program for DNA sequenceanalysis on Apple II microcomputer is described. Use of a binarycoding of nucleotides allows interactive data manipulation ona low-cost configuration with execution times similar to thoseof larger computers. The PEGASE system should prove useful andeasy to use in routine sequence handling and experiment design. Received on July 23, 1984; accepted on August 24, 1984     

A microcomputer program for comparison and alignment of DNA sequence gel readings

During the course of determining the sequence of a large DNAfragment, it is necessary to cross-check numerous gel readingsfrom different DNA fragments, in order to track and eliminatemistakes. An algorithm is presented that takes advantage ofthe high degree of homology between such sequences to constructan alignment of the matching regions. It does not require knowledgeof a starting homology zone, neither large memory areas, evenfor sequences of several kilobases, and it can overcome largegaps or mismatch zones that correspond for instance to misinterpretationof compressions on sequence gels. This algorithm has been implementedin 6502 assembly language on an Apple II computer as an extensionto the PEGASE sequence handling system. Received on May 21, 1985; accepted on July 19, 1985     

Effects of A:T base pairs on the B-Z conformational transitions of DNA

Effects of A:T base pairs on the propensity of B to Z conformational transitions have been investigated by the CD salt titrations on d(CG)5' d(GC)5' terminal or central A:T replaced decamers, and terminal A:T appended dodecamers. The presence of A:T at the center greatly inhibits the B to Z transition of both G:C decamers. Moderate Z inhibitions are shown by terminal A:T replacements and additions to d(CG)5' with the former exhibiting a stronger effect. In contrast, the addition and replacement with A:T at the terminals of d(GC)5 facilitate the B to Z conversion, with the replacement exhibiting a somewhat more pronounced effect. These results may be rationalized in terms of the number of contigous CG sequences present in an oligomer and the relative inhibitory effects of other dinucleotide sequences. Our results also suggest that some short oligomers with purine at the 5'-end, such as d[A(CG)nT] with n greater than or equal to 2, may likely crystallize as Z conformations.     

RecA-promoted sliding of base pairs within DNA repeats: quantitative analysis by a slippage assay

RecA that catalyses efficient homology search and exchange of DNA bases has to effect major transitions in the structure as well as the dynamics of bases within RecA-DNA filament. RecA induces slippage of paired strands in poly(dA)-poly(dT) duplex using the energy of ATP hydrolysis. Here, we have adopted the targeted ligation assay and quantified the strand slippage within a short central cassette of (dA)(4)-(dT)(4) duplex. The design offers a stringent test case for scoring a cross-talk between A residues with those of T that are diagonally placed on the opposite strand at either -3, -2, -1, +1, +2, or +3 pairing frames. As expected, the cross-talk levels in RecA mediated as well as thermally annealed duplexes were maximal in non-diagonal pairing frame (i.e., 0-frame), the levels of which fell off gradually as the frames became more diagonal, i.e., -3<-2<-1 or +3<+2<+1. Interestingly, the level of cross-talk in naked duplexes was intrinsically less efficient in minus frames than their plus frame counterparts. The asymmetry created in naked duplexes by such a disparity between minus versus plus frames was partially obviated by RecA. Moreover, RecA promoted a significantly higher level of cross-talk selectively in -2 and -1 frames, as compared to that in naked DNA, which suggests a model that the elevated cross-talk in RecA filament may be limited to base pairs housed within the same rather than adjacent RecA monomers.     

System for DNA sequencing with resolution of up to 600 base pairs

A system capable of resolving about 500 bases is of interest for sequencing of longer DNA molecules. Studies on further optimization of resolution on DNA sequencing gels were carried out. The effect of physico-chemical properties of gels and buffers on resolution were tested, e.g. ionic strength and pH of buffers, different buffer systems, acrylamide concentration, crosslinker concentration, type of crosslinker, temperature of polymerization, denaturing conditions, gel length and thickness. Tested were as well different running conditions like electric field, gel temperature, dimension of sample slots. Gels 0.1-0.2 mm thick and up to 1.2 m long were cast and tested routinely. Gel lengths of 60-70 cm (for sequencing up to 350-400 bases) to about 100 cm (above 400 bases) are practicable. Little is gained in resolution by increasing the gel length from 1 to 1.2 m. Resolution was improved using 0.1 mm thick gels, at a higher pH value of 8.6-8.8, and molarity increased to 0.2 M. The sequencing pattern in the region of higher bases could be better resolved on a twice-magnified picture of that region on the autoradiogram. With the long gels (70-120 cm), it is advantageous to obtain the sequence overlap by running in parallel gels of different concentrations, without re-application of samples, all loaded at the same time. Buffer chamber for running of two of three gels and thermostating plates up to 1.2 m long were designed. In this way four to six thermostated gels can be run from a power supply with two inputs. Three 1 m long gels (concentrations: 4%, 6%, 12-16%) are loaded with several samples of DNA to be sequenced and run in parallel without re-application of the samples. With good samples, the sequence overlap from the gels could be counted up to 500 base pairs, with exceptionally good samples closer to 600 bases. At present this number seems to be near the limit of the resolving power of the polyacrylamide gels.     

MOLECULAR DESIGNER: an interactive program for the display of protein structure on the IBM-PC

A BASIC interactive graphics program has been developed forthe IBM-PC which utilizes the graphics capabilities of thatcomputer to display and manipulate protein structure from coordinates.Structures may be generated from typed files, or from BrookhavenNational Laboratories' Protein Data Bank data tapes. Once displayed,images may be rotated, translated and expanded to any desiredsize. Figures may be viewed as ball-and-stick or space-fillingmodels. Calculated multiple-point perspective may also be addedto the display. Docking manipulations are possible since morethan a single figure may be displayed and manipulated simultaneously.Further, stereo images and red/blue three-dimensional imagesmay be generated using the accompanying DESIPLOT program andan HP-7475A plotter. A version of the program is also currentlyavailable for the Apple Macintosh. Full implementation on theMacintosh requires 512 K and at least one disk drive. Otherwisethis version is essentially identical to the IBM-PC versiondescribed herein. Received on July 12, 1985; accepted on August 1, 1985     

A Pascal program for weighted least squares regression on a microcomputer

Weighted least-squares regression has been programmed in Pascal for a microcomputer. A double precision Pascal compiler and the Motorola 6809 assembler produce a fast machine-code program occupying 22,000 bytes of memory when appended to the Pascal run-time module. Large data sets fit in the remaining memory. A regression with 72 observations and 24 parameters runs in 7 min, excluding optional print out of large matrices. The maximum dimensions of the design matrix, X, can be altered by modifying two Pascal constants. Minor changes to the Pascal source program will make it compatible with other Pascal compilers. The program optionally orthogonalises the X matrix to detect linearly-dependent columns in X, and/or generate orthogonal parameter estimates. After orthogonalizing X and fitting the model, the parameter estimates for the original X can be retrieved by the program. Regressions on a repeatedly reduced model are performed through elimination of columns in X until the minimum adequate model is obtained.     

Preparation and properties of a nucleosomal particle containing 180 base pairs of DNA

If chromatin from chicken erythrocytes is enzymatically degraded in the presence of histone H5, nucleosomal DNA usually exceeds the size of 140 base pairs. Conditions are derived allowing the isolation of a 180 base pair particle which is subsequently characterized by histone binding and thermal denaturation studies. Association of H5 to such a particle is cooperative and occurs with a larger affinity than binding to specimens in the range of 140– 170 base pairs. Thermal denaduration studies show that some of the extra DNA participates in the main transition at 74°C (1 mm Na-cacodylate) indicating a tight binding to the histone core. Another part of the extra segment is loosely associated with the core but also distinct from free DNA.     

Intercalation of daunomycin into stacked DNA base pairs. DFT study of an anticancer drug

We have computationally studied the intercalation of the antitumor drug daunomycin into six stacks of Watson-Crick DNA base pairs (i.e., AT-AT, AT-TA, GC-AT, CG-TA, GC-GC, GC-CG) using density functional theory (DFT). The proton affinity of the DNA intercalator daunomycin in water was computed to be 159.2 kcal/mol at BP86/TZ2P, which is in line with the experimental observation that daunomycin is protonated under physiological conditions. The intercalation interaction of protonated daunomycin with two stacked DNA base pairs was studied through a hybrid approach in which intercalation is treated at LDA/TZP while the molecular structure of daunomycin and hydrogen-bonded Watson-Crick pairs is computed at BP86/TZ2P. We find that the affinity of the drug for the six considered base pair dimers decreases in the order AT-AT > AT-TA > GC-AT > GC-TA > GC-CG > GC-GC, in excellent agreement with experimental data on the thermodynamics of the interaction between daunomycin and synthetic polynucleotides in aqueous solution. Our analyses show that the overall stability of the intercalation complexes comes mainly from pi-pi stacking but an important contribution to the computed and experimentally observed sequence specificity comes from hydrogen bonding between daunomycin and hetero atoms in the minor groove of AT base pairs.     

Alternative heterocycles for DNA recognition: a 3-pyrazole/pyrrole pair specifies for G.C base pairs

Synthetic ligands comprising three aromatic amino acids, pyrrole (Py), imidazole (Im), and hydroxypyrrole (Hp), specifically recognize predetermined sequences as side-by-side pairs in the minor groove of DNA. To expand the repertoire of aromatic rings that may be utilized for minor groove recognition, three five-membered heterocyclic rings, 3-pyrazolecarboxylic acid (3-Pz), 4-pyrazolecarboxylic acid (4-Pz), and furan-2-carboxylic acid (Fr), were examined at the N-terminus of eight-ring hairpin polyamide ligands. The DNA binding properties of 3-Pz, 4-Pz, and Fr each paired with Py were studied by quantitative DNase I footprinting titrations on a 283 bp DNA restriction fragment containing four 6-bp binding sites 5'-ATNCCTAA-3' (N = G, C, A, or T; 6-bp polyamide binding site is underlined). The pair 3-Pz/Py has increased binding affinity and sequence specificity for G.C bp compared with Im/Py.