The nucleotide sequence of the replication origin beta of the plasmid R6K

We h ave identified by molecular cloning a region of 283 base pairs of the HindIII 2 fragment of R6K which corresponds to the region of the replication origin beta. This 283 base-pair DNA fragment, when present contiguously with the structural gene for the replication initiation protein of R6K, encoded in the HindIII 9-15 and part of HindIII 2 restriction fragments, will support the replication of a plasmid chimera containing the pBR322 replicon in a pol Ats host at the nonpermissive temperature. The nucleotide sequence of the region of replication origin beta has been determined. The nucleotide sequence has some homology with the ori gamma region of R6K; it has a 15-base-pair homology with the replication origin of Escherichia coli.     

Complete nucleotide sequence and gene organization of plasmid NTP16.

We have determined the complete nucleotide sequence of the 8.3-kb multicopy plasmid NTP16 and produced a functional map of its gene organization. Sixty percent of the plasmid DNA comprises transposon-derived sequences; in the remaining 3320 bp, we have identified three protein coding regions. NTP16 has a ColE1-type replication system, a cis-acting stability locus and a mobilization system comprising an oriT site and one mobilization protein. The roles of the other two protein products of this plasmid are unknown, but they are possibly involved in the plasmid incompatibility system.     

The nucleotide sequence surrounding the origin of DNA replication of Col E1.

The DNA of Col E1 replicates from a unique origin located at a distance of 17-19% of the genome length from the single Eco RI clevage site. The nucleotide sequence about this site has been determined by a combination of RNA and DNA sequencing techniques. The principal features of the sequence are two palindromes, one of which resembles a palindrome located in the intercistronic region of 0X174. The sequence also contains stretches of purine and pyrimidine clusters of the following compositions: pAT5G, pC2T5G, pGT5G. The origin sequence demonstrates that initiation of DNA replication takes place in an intercistronic region of Col E1DNA, although the possibility that this region makes small polypeptides 30-40 residues long cannot be strictly eliminated at this time.     

The nucleotide sequence surrounding the replication origin of the cop3 mutant of the bacteriocinogenic plasmid Clo DF13.

The nucleotide sequence from about 100 base-pairs downstream to about 600 base pairs upstream the CloDF13 replication origin has been determined. A comparison of this sequence with the corresponding ColE1 origin sequence reveals that: The sequence at the origin of replication is conserved. There are large differences in the nucleotide sequence downstream the replication origin, whereas there is a large homology in the region of about 410 base-pairs upstream the replication origin. This conserved region might code for a largely homologous basic, arginine rich polypeptide of about 45 amino-acids, for both ColE1 and CloDF13. Although there are large differences in the primary structure of the region coding for the 100 nucleotide RNA, the secondary structure of this region seems to be conserved.     

The nucleotide sequence of the replication control region of the resistance plasmid R1drd-19

Summary The region of plasmid R1 containing the replication control genes has been sequenced using the Maxam-Gilbert method. The nucleotide sequence of two small PstI restriction fragments (a total of about 1,000 base pairs) was determined for the wildtype R1 plasmid as well as for two different copy mutants. It was found that one copy mutant has a single base substitution in the fragment which was recently shown to harbor an important inc/cop gene (Molin and Nordström 1980). Furthermore, the sequence indicates the presence of a structural gene that codes for a polypeptide of size 10,500 daltons. Possible gene products predicted from the nucleotide sequences and their role in replication control are discussed.     

The replication origin of pSC101: the nucleotide sequence and replication functions of the ori region

The nucleotide sequence of a 770-bp ori region of plasmid pSC101 is presented. The sequence shows homologies to some parts of Escherichia coli oriC and phage G4 ori. Several other features are an 80-bp A + T-rich region overlapping a part of the region homologous to oriC, three direct repeats of an 18-bp sequence adjacent to the A + T-rich region, a typical promoter sequence just upstream of the longest open reading frame (ORF) and a long inverted repeat sequence overlapping the putative promoter region. Analysis of successive deletions by BAL31 exonuclease demonstrated that one of the regions homologous to oriC along with the A + T-rich region are essential for autonomous replication of the plasmid. The three 18-bp repeats are responsible for incompatibility phenotype. The region containing the promoter-like sequence is required for expression of a trans-acting function.     

DNA bending near the replication origin of IncFII plasmid NR1.

The DNA replication origin of plasmid NR1 is located approximately 190 base pairs downstream from the 3' end of the repA1 gene, which encodes the essential initiation protein for replication of the plasmid. Restriction endonuclease fragments that contain the NR1 replication origin and its flanking sequences at circularly permuted positions were obtained by digesting oligomers of ori-containing DNA fragments with sets of enzymes that each cut only once in every ori fragment. Polyacrylamide gel electrophoresis of these permuted restriction fragments showed anomalous mobilities, indicating the presence of a DNA bending locus. Through analysis of the relative mobility plots of these permuted fragments, we found one or two possible DNA bending sites located in the intervening region between the repA1 gene and the replication origin of NR1. It seems possible that DNA bending in this region might help to orient the replication origin alongside the repA1 gene, which could contribute to the cis-acting character of the RepA1 initiation protein.     

The replication origin region of Escherichia coli: nucleotide sequence and functional units

The minichromosome pCM959 contains the DNA segment from bp -677 (left) to bp + 3335 (right) of the Escherichia coli replication origin, oriC. The nucleotide sequence of this plasmid was determined. The coding regions for proteins were identified, and the possible function of those proteins is discussed. Within oriC two extended systems of dyad symmetry were found, and their possible significance is considered.     

The nucleotide sequence of the rep gene of cyanobacterial plasmid pSM1

The nucleotide sequence was established for the rep gene of plasmid pSM1 isolated from cyanobacterium Plectonema boryanum CALU 465. Both nucleotide sequence and the encoded amino acid sequences showed 98% homology to the corresponding sequences of small plasmids pPF1, pGL3, pPBS1, pBLX, and pPB1. An active center was identified in the replicative protein sequences.     

Critical sequences in the core of the P1 plasmid replication origin.

The core of the P1 plasmid replication origin consists of a series of 7-bp repeats and a G+C-rich stretch. Methylation of the GATC sequences in the repeats is essential. Forty different single-base mutations in the region were isolated and assayed for origin function. A single-base change within any 7-bp repeat could block the origin, irrespective of whether GATC bases were affected. The repeats themselves were critical, but the short intervals between them were not. Mutations in the G+C-rich region showed it to be a spacer whose exact length is important but whose sequence can vary considerably. It maintains a precise distance between the 7-bp repeats and binding sites for the P1 RepA initiator protein. It may also serve as a clamp to limit strand separation during initiation.     

The Escherichia coli terB sequence affects maintenance of a plasmid with the M13 phage replication origin.

Replication initiated at the bacteriophage M13 origin can be affected by interaction of a properly oriented termination signal terB and the Tus protein. The effect can be alleviated by overproduction of the M13 replication gene protein II.     

The nucleotide sequence of a DNA fragment from the replication origin of the antibiotic resistance factor R1drd19

Summary The recombinant plasmid pRK101 contains a DNA fragment which carries the complete replication origin of the antibiotic resistance factor R1drd-19 inserted into the vector plasmid pBR322. In a spontaneously arising mutant of this plasmid (pRK 103) a deletion of about 215 base pairs (bp) has been detected by heteroduplex analysis and mapping with restriction endonucleases. Essential parts of the replication origin must be located in the deleted sequence. The deletion mutant pRK103, in contrast to its parent plasmid pRK101 is not replicated under the control of the R1 replicon, even when the R1 factor or copy mutants of it are present within the same cell. These latter plasmids can complement a plasmid-specific protein not coded by pRK101 but essential for R1-directed replication. The nucleotide sequence of a 252 bp HpaII fragment covering about 170–200 bp of the deletion was determined. This piece of DNA is rich in G and C and contains a series of small palindromes, symmetrically arranged repeated sequences and short selfcomplementary structures which may be of significance for the initiation of the DNA replication. The possibility that the sequenced DNA fragment comprises a major part of the replication origin of R1drd-19 is discussed.     

Localization of the replication origin of plasmid pE194.

The pE194 replication origin was localized to a 265-base-pair interval by analyzing the ability of purified pE194 restriction fragments to direct replication of heterologous plasmids. Replication was dependent upon RepF protein supplied in trans. The origin region contained a GC-rich dyad symmetry which may serve as the RepF target.     

Analysis of the entire nucleotide sequence of the cryptic plasmid of Chlamydia trachomatis serovar L1. Evidence for involvement in DNA replication.

Chlamydia trachomatis serovar L1/440/LN possesses a 7498bp plasmid which was designated pLGV440. The plasmid was cloned at the BamH1 site of pAT153 into Escherichia coli and the recombinant plasmid was designated pCTL1. A detailed restriction endonuclease map of pCTL1 was constructed. A fragment of the chlamydial plasmid was shown to function as a promoter in E. coli when placed upstream of the lacZ gene. The entire plasmid was sequenced by the chain termination method. Open reading frames were identified from the resulting consensus sequence together with a candidate for the plasmid origin of replication consisting of four perfect tandem repeats of a 22bp sequence, an A:T rich sequence and an open reading frame which could generate a 34.8kdal product. The predicted polypeptide products of the open reading frames were compared by computer with all reported protein sequences. Homology of the predicted polypeptide product of an open reading frame to the E. coli dnaB protein and the analogous product of gene 12 of bacteriophage P22 is described.     

Characterization of the replication origin of the myxobacterial self-replicative plasmid pMF1

Thus far, pMF1 is the only endogenous myxobacterial plasmid whose replication mechanism is unclear. In this study, we determined that the plasmid replicates via the theta-mode. The pMF1.14 gene, located in the pMF1.13-pMF1.15 operon (repABC), encodes an essential replication initiation protein that was predicted to have no typical DNA/protein binding motifs but contains rich disordered regions. The pMF1 replication-related essential cis-acting DNA region, approximate 370bp, was located within pMF1.14, and was found to contain several directly and inverted atypical repeats. The unique characteristics of the pMF1 replicon are suggested to be the reason for its strict narrow host range in Myxococcus cells.