Marker-dependent recombination in T4 bacteriophage. III. Structural prerequisites for marker discrimination.

Abstract Distance- as well as marker-dependence of genetic recombination of bacteriophage T4 was studied in crosses between rIIB mutants with known base sequences. The notion of a "basic recombination," which is the recombination within distances shorter than hybrid DNA length in the absence of mismatch repair and any marker effects, was substantiated. The basic recombination frequency per base pair can serve as an objective parameter (natural constant) of general recombination reflecting its intensity. Comparative studies of the recombination properties of rIIB mutants with various sequence changes in the mutated sites showed that the main factor determining the probability of mismatch repair in recombination heteroduplexes is the length of a continuous heterologous region. A run of A:T pairs immediately adjoining the mismatch appears to stimulate its repair. In the case of mismatches with DNA strands of unequal length, formed by frameshift mutations, the repair is asymmetric, the longer strand (bulge) being preferentially removed. A pathway for mismatch repair including sequential action of endonuclease VII (gp49)----3'----5' exonuclease (gp43)----DNA polymerase (gp43)----DNA ligase (gp30) was proposed. A possible identity of the recombinational mismatch repair mechanism to that operating to produce mutations via sequence conversion is discussed.</jats:p

Genetic recombination in bacteriophage T4: single-burst analysis of cosegregants and evidence in favor of a splice/patch coupling model.

Abstract To reveal the structure of penultimate DNA intermediates in T4 bacteriophage recombination, resolution of which produces free recombinant molecules, a single-burst analysis of the recombinant progeny was made in multifactor crosses, enabling one to determine quantitatively the different recombinants generated by one or two exchanges within the same chromosome segment. It was found that double and single exchanges are highly correlated in T4 recombination. These results were interpreted as evidence for simultaneous formation of a splice/patch pair as the primary recombination products. A recombination model called here the "splice/patch coupling model" is presented according to which resolution of a single DNA intermediate results in two linear heterozygous molecules containing a patch and a splice, respectively, in homologous positions.</jats:p

Marker-dependent recombination in T4 bacteriophage. IV. Recombinational effects of antimutator T4 DNA polymerase.

Abstract Recombinational effects of the antimutator allele tsL42 of gene 43 of phage T4, encoding DNA polymerase, were studied in crosses between rIIB mutants. Recombination under tsL42-restricted conditions differed from the normal one in several respects: (1) basic recombination was enhanced, especially within very short distances; (2) mismatch repair tracts were shortened, while the contribution of mismatch repair to recombination was not changed; (3) marker interference at very short distances was augmented. We infer that the T4 DNA polymerase is directly involved in mismatch repair, performing both excision of a nonmatched single strand (by its 3'--&amp;gt;5' exonuclease) and filling the resulting gap. A pathway for the mismatch repair was substantiated; it includes sequential action of endo VII (gp49)--&amp;gt;3'--&amp;gt;5' exonuclease (gp43)--&amp;gt;DNA polymerase (gp43)--&amp;gt;DNA ligase (gp30). It is argued that the marker interference at very short distances may result from the same sequence of events during the final processing of recombinational intermediates.</jats:p

Double-Strand Break Repair in Bacteriophage T4: Recombination Effects of 3′–5′ Exonuclease Mutations

The role of 3′–5′ exonucleases in double-strand break (DSB)-promoted recombination was studied in crosses of bacteriophage T4, in which DSBs were induced site specifically within the rIIB gene by SegC endonuclease in the DNA of only one of the parents. Frequency of rII(+) recombinants was measured in two-factor crosses of the type i × ets1, where ets1 designates an insertion in the rIIB gene carrying the cleavage site for SegC and i's are rIIB or rIIA point mutations located at various distances (12–2040 bp) from the ets1 site. The frequency/distance relationship was obtained in crosses of the wild-type phage and dexA1 (deficiency in deoxyribonuclease A), D219A (deficiency in the proofreading exonuclease of DNA polymerase), and tsL42 (antimutator allele of DNA polymerase) mutants. In all the mutants, recombinant frequency in crosses with the i-markers located at 12 and 33 bp from ets1 was significantly enhanced, implying better preservation of 3′-terminal sequences at the ends of the broken DNA. The effects of dexA1 and D219A were additive, suggesting an independent action of the corresponding nucleases in the DSB repair pathway. The recombination enhancement in the dexA1 mutant was limited to short distances (<100 bp from ets1), whereas in the D219A mutant a significant enhancement was seen at all the tested distances. From the character of the frequency/distance relationship, it is inferred that the synthesis-dependent strand-annealing pathway may operate in the D219A mutant. The recombination-enhancing effect of the tsL42 mutation could be explained by the hypothesis that the antimutator 43Exo removes a shorter stretch of paired nucleotides than the wild-type enzyme does during hydrolysis of the unpaired terminus in the D-loop intermediate. The role of the proofreading exonuclease in the formation of a robust replicative fork is discussed

Peculiarities of distance learning in higher education: The teacher’s functions as a chat communication organizer

The purpose of this article is to define the functions of the teacher as the organizer of chat communication, which has become one of the foundations of Internet communication. The leading method for the study of this problem is the method of situational modeling, which allows considering chat-communication as a process of new social interaction, requiring the leading role of the teacher-organizer, ensuring the maximum involvement of all participants-communicators in this process in order to assimilate new knowledge by the perceiver (learners). This article presents new methodological methods of teacher's work within the framework of chat-communication, defines the important functions of a teacher, who communicates knowledge in the changed socio-cultural environment. The presented methodological material is designed to help teachers of higher education to use chat-communication as a new channel of knowledge transfer from a teacher to a student to adapt to the new communicative realities, to give awareness of the continuity of the educational process, even in a situation of significant changes in conditions of its implementation.&#x0D;  &#x0D; Keywords: distance learning, higher education, internet communication, chatting, teacher functions</jats:p

MARKER-DEPENDENT RECOMBINATION IN T4 BACTERIOPHAGE. I. OUTLINE OF THE PHENOMENON AND EVIDENCE SUGGESTING A MISMATCH REPAIR MECHANISM

ABSTRACT In standard crosses, some rIIB mutants of T4 phage were found to be susceptible to an extra recombination mechanism to which the other mutants were much less susceptible. The following observations were interpreted as evidence for the mismatch-repair nature of the phenomenon: (1) Marker-dependent recombination generates exclusively double exchanges at both sides of the marker. (2) Marker-dependent recombination is highly sensitive to DNA base sequence at the site of the marker and to that at the corresponding site on the chromosome of the other parent. (3) Within certain limits, the contribution of the marker-dependent mechanism to the total recombination frequency is distance-independent and thus constitutes a constant component.</jats:p