Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
10143495 | Gene | 2018 | 28 Pages |
Abstract
Five open reading frames were predicted based on the nucleotide sequence analysis. The deduced amino acid sequence of all predicted ORFs didn't show any similarity with any known proteins. Three palindroms were detected and two promoter sequences were predicted in both strands. With electron microscopy (TEM) analysis, the replication intermediates were seen as typical Q-shaped molecules that committing pHIG22 replicates via the Theta replication mechanism. A 2012â¯bp region among 387 and 614â¯bp of pHIG22 was determined as minimal replicon which carries the elements necessary for plasmid replication and ori region. Furthermore, quantitative real-time PCR showed that the relative copy number of pHIG22 was estimated to be 148.2â¯Â±â¯4.7 copies per chromosome equivalent. The new Theta type plasmid would be useful and beneficial to build vectors for cloning of thermophilic genes and in vivo protein engineering.
Keywords
dNTPORFIPTGkDaμgμLμMTBEE. coliKCIMgSO4milimolarX-GalHTKrpoBddH2OMin.qPCRμmolAMPDNA°Cdouble distilled waterAmpicillinEDTAethylenediaminotetraacetic aciddeoxyribonucleic acidEscherichia coliAngströmTemBETATris-EDTAReplication originbase pairdaltonDegreeminuteDeltaMelting TemperatureSecondMagnesium sulphateopen reading frameLuria Bertani brothliterUniteKanMolarmillilitermmolmilligramTransmission electron microscopynanomolarnanogrampolymerase chain reactionquantitative polymerase chain reactionPCRKilo baseCryptic plasmidthreshold cycleoptical densityKanamycinMagnesium chloridePotassium chlorideCalcium chlorideKilo Dalton
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Authors
Halil Ibrahim Guler, Esma Ceylan, Sabriye Canakci, Ali Osman Belduz,