Shaaban Mohamed T, Abdelhamid Reham M, Zayed Muhammad, Ali Safaa M
Botany and Microbiology Department, Menoufia University, Shebin El-Kom, Egypt.
Genetic Engineering and Biotechnology Research Institute (GEBRI), City of Scientific Research and Technological Applications, Alexandria, Egypt.
Biotechnol Rep (Amst). 2022 Jan 25;34:e00706. doi: 10.1016/j.btre.2022.e00706. eCollection 2022 Jun.
Diseases and epidemics in the current days need new types of antibiotics in order to be able to eliminate them. The goal of this research is to use metagenomics to identify isolated utilitarian gene (s) as antimicrobial specialists. Collection of diverse locations from sea sediment samples from Alexandria and extraction of total DNA, restriction enzyme fragmentation, cloning into pUC19 vector, and expression of the isolated gene(s) in DH5 were all part of the process. Characterization of Antimicrobial agent was done for the best clone for antimicrobial agent's production to detect efficiency, optimum pH, thermal stability, pH stability, effect of different compounds on antimicrobial activity, and residual activity of product after preservation in room temperature. Amino acid sequence of RSMM C3 gene (1250 bp) was 72% identity with sp. The ideal temperature level of the RSMM C3 antimicrobial agent production was 36 °C. The antimicrobial agent RSMM C3's stability was stable at -20 °Celsius for up to two months without thawing. The antibacterial agent RSMM C3 was stable at 4 °C for 14 days without loss in activity. The ideal pH level of the RSMM C3 antimicrobial agent was 6. Remain activity was gradually decreased at pH 5, 6, 6.5 and 7 (86.1, 96.9, 97.2 and 94.9%, respectively). On the other hand, residual activity was (92 and 84%) at (pH 7.5 and 8) for 8 days. The tested antimicrobial RSMM C3 was stable against 1 mM of different compounds (DMSO, Glycerol, NaCl, CaCl, MgCl, ZnCl, FeSO, MnSO and CuSO). The research provides for the Metagenomics technique that has the ability for the production of novel antimicrobial agents produced by clone RSMM C3 which has a wide spectrum activity towards different microorganisms comparing to other antibiotics as Ampicillin and Tetracycline.
当今的疾病和流行病需要新型抗生素才能将其消灭。本研究的目标是利用宏基因组学来鉴定分离出的实用基因作为抗菌专家。从亚历山大港的海洋沉积物样本中收集不同地点的样本，提取总DNA，进行限制性酶切片段化，克隆到pUC19载体中，并在DH5中表达分离出的基因，这些都是该过程的一部分。对生产抗菌剂的最佳克隆进行抗菌剂特性分析，以检测其效率、最佳pH值、热稳定性、pH稳定性、不同化合物对抗菌活性的影响以及室温保存后产品的残留活性。RSMM C3基因（1250 bp）的氨基酸序列与 种的同一性为72%。RSMM C3抗菌剂生产的理想温度水平为36℃。抗菌剂RSMM C3在-20℃下可稳定保存长达两个月而不融化。抗菌剂RSMM C3在4℃下可稳定保存14天而活性无损失。RSMM C3抗菌剂的理想pH值为6。在pH 5、6、6.5和7时残留活性逐渐降低（分别为86.1%、96.9%、97.2%和94.9%）。另一方面，在pH 7.5和8时，8天后残留活性分别为92%和84%。测试的抗菌剂RSMM C3对1 mM的不同化合物（二甲基亚砜、甘油、氯化钠、氯化钙、氯化镁、氯化锌、硫酸亚铁、硫酸锰和硫酸铜）稳定。该研究提供了宏基因组学技术，该技术有能力生产由克隆RSMM C3产生的新型抗菌剂，与氨苄青霉素和四环素等其他抗生素相比，其对不同微生物具有广泛的活性。
