Tetrad analysis

Johnston, J.R., Baccari, C., and Mortimer, R.K. 2000. Genotypic characterization of strains of commercial wine yeasts by tetrad analysis. Res. Microbiol. 151, 583-590. 

If a gene is nuclear, then a diploid carrying two alleles of the gene will produce by meiosis four spores, two of which will carry one allele, and two the other. Thus, tetrad analysis is said to yield 2 2 segregation for nuclear genes. If a genetic determinant is cytoplasmic, and many copies are present, then when spores are formed cytoplasmic elements will be encased in each spore coat. Thus, in the case of cytoplasmic inheritance, tetrad analysis will usually show 4 0 segregation. 

Fig. 16.9 The genetic map of S. cerevisiae [74]. Centromeres are represented as circles, the left arm of each chromosome being drawn above the centromere. Dashed and dotted lines represent linkages established other than by tetrad analysis. Parentheses enclose genes where exact orientation is unknown. (Published by kind permission of Dr R. K. Mortimer.)...

Tetrad analysis on only partially digested cultures requires little time and has the advantage of reliability. Digestion has to be reduced to a limit which allows a relatively easy bursting of asci with a micromanipulator needle on an agar slab (3% agar in a 0.4% solution of potassium acetate). 

The first point of the stereochemical analysis is in the recognition of the sequence to which a given nucleus is sensitive the problem seems rather obvious for vinyl or vinylidene polymers where the sequence must extend equally from the two sides of the nucleus in question but for diene polymers or those containing heteroatoms, the problem is not so simple. In the present case, the methylene protons are sensitive to the structure of the even sequences, dyads and tetrads, whereas the methyl protons are sensitive to the odd sequences, triads, and pentads. 

To prove the correctness of this analysis we shall try to look for alternative explanations. A stereoregular heterotactic polymer. . . mrmrmr. . . could also explain the presence of a methyl singlet and a well-separated doublet of doublets for methylene protons, in agreement with the spectrum of the first sample in fact, there would be present the mr triad and the rmr tetrad with diastereotopic protons. However, in addition, an equal amount of the mrm tetrad should be present, yet the corresponding singlet is not visible with the required intensity. In the same way other hypothetical structures can be rejected. 

Analysis of the poly(methyl methacrylate) sequences obtained by anionic polymerization was undertaken at the tetrad level in terms of two different schemes (10) one, a second-order Markov distribution (with four independent conditional probabilities, Pmmr Pmrr, Pmr Prrr) (44), the other, a two-state mechanism proposed by Coleman and Fox (122). In this latter scheme one supposes that the chain end may exist in two (or more) different states, depending on the different solvation of the ion pair, each state exerting a specific stereochemical control. A dynamic equilibrium exists between the different states so that the growing chain shows the effects of one or the other mechanism in successive segments. The deviation of the experimental data from the distribution calculated using either model is, however, very small, below experimental error, and, therefore, it is not possible to make a choice between the two models on the basis of statistical criteria only. 

High magnetic fields and in particular C-NMR spectroscopy allow the analysis of even longer configurational sequences (tetrads up to nonads). This proved to be important in particular for the analysis of polyolefins like polypropylene or cycloolefin copolymers (COC). These polymers are available via transition-metal mediated (Ziegler-Natta, metallocene) insertion polymerizations, and the configurational analysis provides deep insight into the respective polymerization mechanisms as well as into the structure-property relationships. 

More convenient and entirely sufficient for the present purpose is the calculation of ratio of rate constants. The calculation will be reviewed for a one-way first-order Markov process. A one-way mechanism is chosen because it is intuitively the most appropriate model for a free radical mechanism. Furthermore it has some experimental support. The assumption of a first-order Markov process does not rule out higher Markov processes. The differentiation between a first-order Markov process and higher order Markov processes is however possible experimentally in very rare cases because it involves the determination of tetrad, pentad, etc. fractions (11, 12, 13, 14). A Bemoullian process is ruled out by the analysis of the data of Table I. 

The PLA tacticity (and stereocontrol mechanism) can be assessed by analysis of the NMR spectrum of the PLA. This can be carried out either by analysis of the homonuclear NMR spectrum, at the tetrad level, or by analysis of the I3C H NMR spectrum (carbonyl region). An illustration showing the assignment of the tetrads in the methane region of the homonuclear decoupled 1H H NMR spectrum is included in Fig. 8. 

It has been recognized that the stabilization by interactions with metal ions is essential in the formation of tetraplex structures. Based on the electrostatic potential analysis of the tetrads, a number of tetrads are suggested as the possible hosts of cations. The interaction between the cation and the tetrads remarkably alter the bonding patterns of the tetrads. 

J. Gu et al., H-bonding patterns in the platinated guanine-cytosine base pair and guanine-cytosine-guanine-cytosine base tetrad An electron density deformation analysis and aim study. J. Am. Chem. Soc. 126, 12651-12660 (2004)... 

Methyl methacrylate was synthesized under different polymerization conditions to give two samples of PMMA (denoted I and II in Table 7-8) that were synthesized under different polymerization conditions. An NMR analysis of these samples gave the following data regarding triad and tetrad sequences. 

---