The transcription machinery of archaea can be roughly classified as a simplified version of eukaryotic organisms. The basal transcription factor machinery binds to the TATA box found around 28 nucleotides upstream of the transcription start site; however, some transcription units lack a clear TATA box and still have TBP/TFB binding over them. This apparent absence of conserved sequences could be a consequence of sequence divergence associated with the upstream region, operon, and gene organization. Furthermore, earlier studies have found that a structural analysis gains more information compared with a simple sequence inspection. In this work, we evaluated and coded 3630 archaeal promoter sequences of three organisms, Haloferax volcanii, Thermococcus kodakarensis, and Sulfolobus solfataricus into DNA duplex stability, enthalpy, curvature, and bendability parameters. We also split our dataset into conserved TATA and degenerated TATA promoters to identify differences among these two classes of promoters. The structural analysis reveals variations in archaeal promoter architecture, that is, a distinctive signal is observed in the TFB, TBP, and TFE binding sites independently of these being TATA‐conserved or TATA‐degenerated. In addition, the promoter encountering method was validated with upstream regions of 13 other archaea, suggesting that there might be promoter sequences among them. Therefore, we suggest a novel method for locating promoters within the genome of archaea based on DNA energetic/structural features.

Background Archaea are a vast and unexplored domain. Bioinformatic techniques might enlighten the path to a higher quality genome annotation in varied organisms. Promoter sequences of archaea have the action of a plethora of proteins upon it. The conservation found in a structural level of the binding site of proteins such as TBP, TFB, and TFE aids RNAP-DNA stabilization and makes the archaeal promoter prone to be explored by statistical and machine learning techniques. Results and discussions In this study, experimentally verified promoter sequences of the organisms Haloferax volcanii, Sulfolobus solfataricus, and Thermococcus kodakarensis were converted into DNA duplex stability attributes (i.e. numerical variables) and were classified through Artificial Neural Networks and an in-house statistical method of classification, being tested with three forms of controls. The recognition of these promoters enabled its use to validate unannotated promoter sequences in other organisms. As a result, the binding site of basal transcription factors was located through a DNA duplex stability codification. Additionally, the classification presented satisfactory results (above 90%) among varied levels of control. Concluding remarks The classification models were employed to perform genomic annotation into the archaea Aciduliprofundum boonei and Thermofilum pendens, from which potential promoters have been identified and uploaded into public repositories.

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Archaea are a vast and unexplored cellular domain that thrive in a high diversity of environments, having central roles in processes mediating global carbon and nutrient fluxes. For these organisms to balance their metabolism, the appropriate regulation of their gene expression is essential. A key momentum in regulating genes responsible for the life maintenance of archaea is when transcription factor proteins bind to the promoter element. This DNA segment is conserved, which enables its exploration by machine learning techniques. Here, we trained and tested a support vector machine with 3935 known archaeal promoter sequences. All promoter sequences were coded into DNA Duplex Stability. After, we performed a model interpretation task to map the decision pattern of the classification procedure. We also used a dataset of known-promoter sequences for validation. Our results showed that an AT rich region around position − 27 upstream (relative to the start TSS) is the most conserved in the analyzed organisms. In addition, we were able to identify the BRE element (− 33), the PPE (at − 10) and a position at + 3, that provides a more understandable picture of how promoters are organized in all the archaeal organisms. Finally, we used the interpreted model to identify potential promoter sequences of 135 unannotated organisms, delivering regulatory regions annotation of archaea in a scale never accomplished before (https://pcyt.unam.mx/gene-regulation/). We consider that this approach will be useful to understand how gene regulation is achieved in other organisms apart from the already established transcription factor binding sites.

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For enumerating viable bacteria, traditional dilution plating to count colony forming units (CFU) has always been the preferred method in microbiology owing to its simplicity, albeit laborious and time-consuming. Similar CFU counts can be obtained by quantifying growing microcolonies in conjunction with the perks of a microscope. Here, we employed a simple method of five microliter spotting of differently diluted bacterial culture multiples times on a single petri plate followed by finding out CFU by counting microcolonies using phase contrast microscope. In this method within four-hour period CFU of an Escherichia coli culture can be found out. Further, within ten-hour period, CFU in a culture of Ralstonia solanacearum, a bacterium with generation time around 3 h, can be estimated. The CFU number determined by microcolonies observed is comparable with that obtained by the dilution plating method. Microcolonies number observed in the early hours of growth (2 h in case of E. coli and 8 h in case of R. solanacearum) were found to remain consistent at later h, though there was noticeable increase in size of the colonies. It suggested that microcolonies observed in the early hours indeed represent the bacterial number in the culture. Though manual, the method is less labor intensive apart from being simple, easy, economical and rapid.

Nucleoid‐associated proteins (NAPs) maintain bacterial nucleoid configuration through their architectural properties of DNA bending, wrapping, and bridging. However, the contribution of DNA structural alterations to DNA‐NAP recognition at the genomic scale remains unresolved. Present work dissects the DNA sequence, shape and altered structural preferences at a genomic scale for six NAPs in Mycobacterium tuberculosis. Results suggest narrower minor groove width (MGW) and higher DNA rigidity are marked for the binding sites of EspR and Lsr2, while mIHF, MtHU and NapM have heterogeneous DNA structural predilections. In contrast, WhiB4–DNA‐binding sites were characterized by wider MGW, highly deformable and less curved DNA. This work provides systematic insight into NAP‐mediated genome organization as a function of DNA structural features.

Motivation Archaea are a vast and unexplored cellular domain that thrive in a high diversity of environments, having central roles in processes mediating global carbon and nutrient fluxes. For these organisms to balance their metabolism, the appropriate regulation of their gene expression is essential. A key momentum in regulating genes responsible for the life maintenance of archaea is when transcription factor proteins bind to the promoter element. This DNA segment is conserved, which enables its exploration by machine learning techniques. Results Here, we trained and tested a support vector machine with 3935 known archaeal promoter sequences. All promoter sequences were coded into DNA Duplex Stability. After, we performed a model interpretation task to map the decision pattern of the classification procedure. We also used a validation dataset of known-promoter sequences. Our results showed that an AT rich region around position − 27 upstream (relative to the start TSS is the most conserved in the analyzed organisms. In addition, we were able to identify the BRE element (-33), the PPE (at -10) and a position at + 3, that provides a more understandable picture of how promoters are organized in all the archaeal organisms. Finally, we used the interpreted model to identify potential promoter sequences of 135 unannotated organisms, delivering regulatory regions annotation of archaea in a scale never accomplished before (https://pcyt.unam.mx/gene-regulation/). We consider that this approach will be useful to understand how gene regulation is achieved in other organisms apart from the already established transcription factor binding sites.