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Biomedical subjects

Iu A Berlin

Publications and source records attributed to Iu A Berlin.

17 recordsLinked to original sources
PubMed1992

[Artificial DNA splicing using directed ligation].

An approach to the directed genetic recombination in vitro has been devised, which allows for joining, in a predetermined chemical-enzymatic way, a series of DNA segments to give a precisely spliced polynucleotide sequence (DNA Splicing by Directed Ligation, SDL). The approach makes use of amplification, by several polymerase chain reactions (PCR), of the chosen DNA segments. The corresponding primers contain recognition sites of the class IIS restriction endonucleases, yielding protruding ends of unique primary structures. The protruding ends of the segments to be joined together are structurally predetermined to make them mutually complementary. Ligation of the mixture of the segments so synthesized gives the desired sequence in an unambiguous way. The suggested approach has been exemplified by the synthesis of a totally processed (intronless) gene encoding human mature interleukin-1 alpha.

Base Sequence↗
PubMed1991

[Allele polymorphism of the DNA loci MET, D7S8, D7S23, linked to the cystic fibrosis gene in some populations of the USSR, in high risk families and in cystic fibrosis patients].

RELP analysis of DNA loci MET, D7S8 and D7S23 was carried out in Leningrad population and partially in populations of Moscow, Azerbaijan, Ukraine, Buryatia as well as in individuals from high risk families and in cystic fibrosis (CF) patients by means of blot hybridization and polymerase chain reaction. Allelic polymorphism of all loci studied in these three groups was found to be quite similar to that in the North-Western Europe and in whites of the North America. Linkage disequilibrium of the alleles studied with the CF gene was especially pronounced for alleles of the D7S23 locus and gradually decreases from KM-19 through CS-7 to XV-2c DNA probes. The data witness genetic homogeneity of the CF mutation in European populations of the USSR and its similarity to this mutation in Western Europe. The significance of these data for potential diagnosis of CF and for heterozygous carrier detection is discussed.

Alleles↗
PubMed1990

[Synthesis of the mature human interleukin-1(alpha) gene by amplification of an mRNA-cDNA duplex in vitro].

Mixture of polyadenylated mRNAs from human monocytes has been subjected to the reverse transcription specifically initiated from the mRNA encoding interleukin 1 alpha by a synthetic polynucleotide complementary to the mRNA's coding 3'-end to yield the corresponding mRNA-cDNA duplex. Under conditions of the polymerase chain reaction, with the above polynucleotide as a downstream primer and an upstream primer corresponding to the beginning of the mature interleukin 1 alpha (AA 113-271) gene, the mRNA-cDNA duplex yielded the desired gene, whose structure was proved by the restriction and sequence analyses. The gene, containing translation initiation and termination triplets, can be used for producing interleukin 1 alpha in various expression systems and as a probe in studies of the lymphokine's biosynthesis. This method of the gene synthesis does not need construction and analysis of cDNA libraries nor synthesis of double-stranded DNA, and can, in principle, make use of the total (non-fractionated) cellular RNA.

Base Sequence↗
PubMed1989

[Structure of the recombination zone during abnormal excision of the transducing bacteriophage lambda plac9].

Investigating molecular mechanism of illegitimate recombinations in prokaryote we study transducing bacteriophages of the lambda lac series. We have carried out physical mapping of bacteriophage lambda plac9 DNA and, by comparing the obtained results with the data on the structure of lambda DNA and lac operon of E. coli, located the phage-bacterial junction corresponding to the lambda-lac9 abnormal excision and elucidated the nucleotide sequence around the junction. It led to the primary structure of phage and bacterial segments in the lysogenic bacterium which took part in the recombinational act leading to the abnormal excision and lambda lac9 formation. Structural homology of the partners in the lambda plac9 excision proved to be lower than in case of the earlier studied lambda plac5 and lambda plac10 whose excision proceeded regioselectively. Various aspects of the crossover area, including the crossover point's probable position and enzymic systems participating in the abnormal excision, are discussed.

Bacteriophage lambda↗
PubMed1988

[Synthetic oligodeoxynucleotides in the study of restriction endonuclease MspI].

Interaction of MspI restriction endonuclease with a series of oligodeoxynucleotides, varying in stability of secondary structure and in location of the restriction site, has been studied. It is shown that a functionally active MspI-site must be double-stranded and flanked from both sides. Separate MspI-cleavage of dodecanucleotides dCGACCCGGGATC and dGATCCCGGGTCG is inhibited by the reaction products as well as by non-homological hexanucleotides dGGTACC and dGGATCC (but not by dCGGCGC). Polyethylene glycol in low concentrations (1-3%) promotes and in higher concentrations (7-14%) inhibits the cleavage. A scheme of MspI functioning is suggested including enzyme's step-by-step recognition of the restriction site and its nonspecific interaction with flanking segments of DNA, which leads to formation of the productive complex.

Base Sequence↗
PubMed1986

[Isolation and properties of recombinant DNA from a plasmid and filamentous phage].

In the course of studying extrachromosomal DNA with composite replicons, a hybrid has been constructed by the in vitro recombination of the filamentous phage M13mp2 DNA (RF) and plasmid pUR222 (ApR). Both parental DNAs contain a fragment of lac-operon (ca. 800 bp), which includes the distal end of lacI gene, lacPO segments, and the lacZ gene proximal region coding for 145 N-terminal amino acid residues of beta-galactosidase and thus providing for alpha-complementation, the effect being cancelled with a polynucleotide insertion at the unique EcoRI site in the lacZ gene segment. E. coli BMH71-18 cells were transformed with the ligated mixture of EcoRI restricts of both DNAs. A phage-like nucleoprotein was isolated from colourless plaques (on the Xgal- and IPTG-supplemented medium); its deproteinization yielded a DNA which contains the ApR-determinant and, according to PAGE, structurally specific staining, restriction analysis, sequencing by the Sanger procedure, and electron microscopy data, is a linear double-stranded molecule comprising the phage and plasmid genomes in an equimolar ratio. Since the hybrid DNA does not display the alpha-complementation effect, both bacterial inserts are in the opposite orientation. Transformation of both phage (F+) and plasmid (F-) hosts with the hybrid DNA led to cultures which, after precipitation of the nucleoprotein from the extracellular medium and deproteinization, afforded the same composite DNA.(ABSTRACT TRUNCATED AT 250 WORDS)

Chromosome Mapping↗
PubMed1986

[Characteristics of the interaction of restriction endonuclease BamHI with oligodeoxynucleotides].

Interaction of the restriction endonuclease BamHI with a series of synthetic oligodeoxynucleotides containing the restriction site has been studied. The enzyme is shown to specifically cut the BamHI site in hexanucleotide (I) and in non-selfcomplementary deca- and octanucleotides (II)-(IV). The data obtained led to the conclusion that BamHI reacts with duplex structures, while playing an important role in their stabilization. In 14-mer (V) BamHI cuts a non-standard half-site GAA to yield the 5'-terminal tri- (rather than hepta-) nucleotide. Hypothetical mechanisms of the process are discussed basing on conception of the role of higher DNA structures in the interaction with restriction endonucleases.

Base Sequence↗
PubMed1986

[Complete primary structure of DNA from the transducing bacteriophage lambda plac5].

In studying molecular mechanisms of specialized transduction, primary structure of the junction between the E. coli gene lacI and the lambda phage locus Ea47 in transducing bacteriophage lambda plac5 has been established. Along with the lambda DNA and E. coli lac operon structures as well as with our earlier data on another phage-bacterial junction in lambda plac5, it lead to the complete sequence of lambda plac5 DNA, including the lac5 substitution, a wellknown segment of lambdoid cloning vehicles. The lambda plac5 DNA is shown to consist of 48645 b.p. distributed as follows: 19368 (lambda left arm) + 3924 (lac5 substitution) + 25353 (lambda right arm). The presence of the phage pbL promoter near to the right end of the lac5 insert is shown. The lacI gene distal end in lambda plac5 proved to be considerably more long-stretched than it used to be believed, coding for 224 C-terminal amino-acid residues of lac repressor. The recombination studied in this paper, similarly to the abnormal prophage excision, occurred near to a Chi-like structure, which is partly homologous to the chi+lacZ site present in lambda plac5. On the basis of the data obtained, a key role of the E. coli RecBC system and Chi sites in the formation of long-stretched deletions in the bacterial cell has been suggested.

Bacteriophage lambda↗
PubMedDate not supplied

[Complex characteristics of the alterations of oncogenes HER-2/ERBB-2, HER-1/ERBB-1, HRAS-1, C-MYC and antioncogenes p53, RB1, as well as deletions of loci of chromosome 17 in colon carcinoma].

Abnormalities of some oncogenes, antioncogenes and losses of heterozygosity (LOH) of chromosome 11p, 17p, and 17q in colorectal carcinomas (CC) was studied. Amplification of ERBB-1/HER-1 oncogene was detected in 2 of 56 cases; ERBB-2/HER-2- in 4 of 62. There was a lack of evidence for C-MYC oncogene amplification (67 cases). LOH of chromosome 11p (HRAS-1 probe) was found in 2 of 37 informative (heterozygous) cases; such events were not accompanied by point mutations in "hot" codons (12th or 61st) in the remaining allele. Prevalence of A3 and A4 alleles of HRAS-1 oncogene (68 cases) as compared to healthy donors was noted. RB-1 (41 cases) and p53 (62 cases) suppressor genes did not show any alterations in Southern-blot analysis. LOH of chromosome 17p (YNZ-22 probe) was found in 15 of 26 heterozygous CC; 17q (THH-59 probe)--in 4 of 16. Analysis of 175th codon of p53 gene revealed only one case of mutation in 35 CC studied. Finally, we were able to detect genetic alterations in 23 of 40 (58%) CC, that were studied on each parameter using Southern-blot. We failed to find any correlation between various molecular abnormalities or clinical characteristics. The data obtained are in disagreement with the view concerning frequent involvement of p53 antioncogene in chromosome 17p deletions.

Base Sequence↗