Direct amplification of new cellulase genes from woodland soil purified DNA

Abstract

Eight genes encoding cellulolytic enzymes were obtained by direct PCR amplification of genomic DNA recovered from woodland soil samples. The direct amplifications were carried out by using primers designed from available online cellulase nucleotide sequences. The isolated genes were all different from each other and homologous to endo-β-1,4-glucanases of Bacillus subtilis. The cellulases were functionally expressed in Escherichia coli and tested on soluble substrate at 37 and 60 °C, showing different cellulolytic activities. Among these, the enzyme renamed CelWS6 exhibited good activity at higher temperatures. Further analysis of CelWS6 showed a high performance in acid environments (between pH 4.0 and 6.0) and at elevated temperatures with its maximum activity at pH 5.0 and 50 °C. At the optimum pH, it was very stable since more than 80 % of its original activity was maintained after an incubation of 120 min at 60 °C. Because the cellulases had different cellulolytic activities, but similar amino acid sequences, it was possible to assess the relationship between sequence and protein function.

The Bacterial Community Structure and Microbial Activity in a Traditional Organic Milpa Farming System Under Different Soil Moisture Conditions

“…The response of the remaining 12, e.g., Bacillus , Flavisolibacter, Pseudomonas , and Streptomyces , to agricultural practices depended on the soil water content. Some of these bacterial groups have been reported to behave as copiotrophs and participate in the degradation of organic material, e.g., Bacillus and Streptomyces ( Garbeva et al, 2003 ; Cucurachi et al, 2013 ; Dang-Thuan et al, 2013 ; Book et al, 2014 ) or compost formation, e.g., Flavisolibacter ( Partanen et al, 2010 ). This confirms once again that bacterial populations are defined by interactions of different factors and a condition that favors a bacterial group, e.g., organic rich milpa soil, alters when a condition changes, i.e., soil water content.…”

Effects of Elevated Tetracycline Concentrations on Aerobic Composting of Human Feces: Composting Behavior and Microbial Community Succession

“…The genus Bacillus is a widespread genus of Firmicutes in feces composting [39] and accounted for 46.41% of the total bacteria in TC500. The genus Bacillus are well known degraders of organic residue, show cellulolytic activity [44], have been found during various stages of composting and are known to produce proteases, chitinases and cellulases [45]. The dominant species at the temperature-fall stage were mainly composed of Sphingobacterium, Moheibacter, Myroides and Pusillimonas.…”

Inducible cellulase production from an organic solvent tolerant Bacillus sp. SV1 and evolutionary divergence of endoglucanase in different species of the genus Bacillus

“…Two pair of primers were used, pair 1: BsFw (forward primer: 5′-GGAATTCCATATGAAACGGTCAATCTC-3′) and BsRv (reverse primer: 5′-TGCGGCCGCCTAATTTGGTTCTGTTCCC-3), 31 and pair 2: CelBF (forward primer: 5′-CCATGGATCATGAGGATGTGAAAACTC-3′) and CelBR (reverse primer: 5′-CTCGAGTGAATTGGTTGTCTGAGCTG-3′). 32 The PCR reactions were carried out in a final volume of 25 μL containing genomic DNA (2 μL), 10× NH 4 buffer (2.5 μL), 50 mM MgCl 2 (2 μL), 10 mM dNTPs (1 μL), 10 mM of each forward/reverse primer (1 μL each), 10 U taq DNA polymerase (0.3 μL) and sterile double distilled water (15.2 μL).…”