Studies of novel transducing variants of lambda: Dispensability of genes N and Q

Court, Donald L.; Sato, Koki

doi:10.1016/0042-6822(69)90060-9

Cited by 171 publications

(46 citation statements)

References 12 publications

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“…Transcription on the left passes through the N gene and, on the right the cro gene, before encountering Rho-dependent terminators tL1 and tR1 respectively. Approximately 50% of the transcripts initiating at pR terminate at tR1, with the remainder (Court and Sato, 1969) is defined by a 2.8 kb deletion (Fiandt et al, 1971) that removes three regions of transcription termination (Cheng et aL, 1995). The following lists the activities of the products of genes not discussed in detail in the text: repressors, cl and cro; transcription activator, cll; replication, O and P. The position of regulatory signals are shown below the listing of genes: small open boxes, terminators; large filled boxes, nut sites; open circles, promoters.…”

Section: Prologuementioning

confidence: 99%

Transcription antitermination: the λ paradigm updated

Friedman

Court²

1995

Molecular Microbiology

128

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Section: Prologuementioning

confidence: 99%

Transcription antitermination: the λ paradigm updated

Friedman

Court²

1995

Molecular Microbiology

128

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“…The presence of Sal128 and Las971 allowed for amplification of the recombinant in subsequent cycles. The 840-bp fragment in which the 26 bp of S. typhimurium DNA was replaced with 11 bp of E. coli DNA was digested with PvuI and used to replace the analogous fragment of the nusA st gene in pNAS1000. The resulting plasmid, pHSE19, was sequenced between the PvuI restriction sites of the DNA insert to confirm that the inserted fragment had the desired deletion-substitution and that no additional changes were introduced by the PCR and cloning steps.…”

Section: Methodsmentioning

confidence: 99%

“…Early A transcription initiating at promoters PL and PR partially terminates at terminators tLl (15, 70) and tRl (10), respectively. The escaping transcripts in the latter case terminate completely at a collection of terminators in the nin region (8,9,11,48, 52) (Fig. …”

mentioning

confidence: 99%

Escherichia coli-Salmonella typhimurium hybrid nusA genes: identification of a short motif required for action of the lambda N transcription antitermination protein

et al. 1994

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The Escherichia coli nusA gene, nusA,E, encodes an essential protein that influences transcription elongation.Derivatives of E. coli in which the SalmoneUla typhimurium nusA gene, nusAs, has replaced nusA$4 are viable. The nusA gene of Escherichia coli encodes an essential 54,400-Mr protein that functions in transcription elongation (reviewed in references 20, 29, and 83) and termination (16,37,49,75,81). Consistent with these roles, NusA has been shown to influence pausing of RNA polymerase (16,17,47,50,75,76,85). The demonstration of an association between NusA and RNA polymerase (31,35,43,46,74) suggests that NusA may influence transcription by a direct interaction with polymerase.NusA was identified through its role in the regulated expression of phage X genes (reviewed in references 13, 20, 23, and 67). Early A transcription initiating at promoters PL and PR partially terminates at terminators tLl (15, 70) and tRl (10), respectively. The escaping transcripts in the latter case terminate completely at a collection of terminators in the nin region (8,9,11,48, 52) (Fig.

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“…The right arm carries a deletion (32) Alint-cIIII originally made in vitro by removing DNA from between the two BamHI sites at 580 and 714 by M. Gottesman, the KH54 deletion (33) removing the rex and cI genes, and the nin 5 deletion (34). The bar underneath the restriction map of X1059 indicates the region of the phage removed in genetic manipulation experiments.…”

Section: Growthmentioning

confidence: 99%

Novel bacteriophage lambda cloning vector.

Karn

Brenner

Barnett

et al. 1980

Proc. Natl. Acad. Sci. U.S.A.

359

128

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A simple method for generating phage collections representing eukaryotic genomes has been developed by using a novel bacteriophage A vector, X1059. The phage is a BamHI substitution vector that accommodates DNA fragments 6-24 kilobases long. Production of recombinants in X1059 requires deletion of the X red and gamma genes. The recombinants are therefore spiP and may be separated from the spi+ vector phages by plating on strains lysogenic for bacteriophage Pg. Random fragments suitable for insertion into X1059 are obtained by partial digestion of high molecular weight eukaryotic DNA with Sau3a. This restriction enzyme cleaves at the sequence G-A-T-C and leaves a 5'-tetranucleotide "sticky end." Because GA-T-C extensions are also produced by BamHI cleavage, these fragments may be annealed directly to BamHI-cleaved X1059. By using these methods, a set of clones covering the entire Caenorhabditis elegans genome was constructed. DNA segments which include the unc-54 myosin heavy chain gene have been isolated from this collection. Fractionation of genomes was an intractable problem until the introduction of recombinant DNA techniques. These methods eliminate the necessity for physical separations of DNA segments and permit the isolation of structural genes from collections of randomly cloned DNA fragments (1-10).Given a suitable probe, any eukaryotic gene may be isolated from a pool of cloned fragments, provided that it is large enough to give sequence representation of an entire genome. The simple multicellular eukaryote Caenorhabditis elegans has a haploid DNA content of approximately 8 X 107 base pairs (bp) (11). If random DNA cleavage and uniform cloning efficiency are assumed, a collection of 8 X 104 clones with an average length of 104 bp will be sufficient to include any genomic sequence with >99% probability. Similarly, the human genome with 2 X 109 bp will be covered by 106 clones 104 bp long.We have developed a novel bacteriophage X cloning vector, X1059, with properties that simplify the construction of recombinant phage collections representing genomes. The strategy is shown in Fig. 1 The structure of A1059 is shown in Fig. 2. The phage is a BamHI substitution vector composed of three BamHI fragments: a 19.6-kb left arm carrying the genes for the A head and tail proteins, a 17-kb central fragment, and a 9.4-kb right arm carrying the A replication and lysis genes. The two arms of the vector contain all the essential functions required for A replication and maturation in a DNA sequence 58.2% of the wildtype length. Viable phages are produced when these arms are hybridized with internal DNA fragments 12.8% to 49.8% the length of wild-type A (6.3-24.4 kb). The two arms alone do not produce viable phages, because lambdoid phages require genome sizes between 70% and 108% of the wild-type DNA to fill the phage heads properly (37, 38).The central BamHI fragment of the vector balancing the phage arms carried the A red (exo and 3 genes) and gamma functions under the control of the leftward promoter (pL) an...

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Studies of novel transducing variants of lambda: Dispensability of genes N and Q

Cited by 171 publications

References 12 publications

Transcription antitermination: the λ paradigm updated

Transcription antitermination: the λ paradigm updated

Escherichia coli-Salmonella typhimurium hybrid nusA genes: identification of a short motif required for action of the lambda N transcription antitermination protein

Novel bacteriophage lambda cloning vector.

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