Molecular Biology Research Communications
Volume:8 Issue: 4, Dec 2019

Recent advances in DNA sequencing techniques have led to an increase in the identification of single nucleotide polymorphisms (SNPs) in BRCA1 and BRCA2 genes, but no further information regarding the deleterious probability of many of them is available (Variants of Unknown Significance/VUS). As a result, in the current study, different sequence- and structure-based computational tools including SIFT, PolyPhen2, PANTHER, SNPs&GO, FATHMM, SNAP, PhD-SNP, Align-GVGD, and I-Mutant were utilized for determining how resulted BRCA protein is affected by corresponding missense mutations. FoldX was used to estimate mutational effects on the structural stabilityof BRCA proteins. Variants were considered extremely deleterious only when all tools predicted them to be deleterious. A total of 10 VUSs in BRCA1 (Cys39Ser, Cys64Gly, Phe861Cys, Arg1699Pro, Trp1718Cys, Phe1761Ser, Gly1788Asp, Val1804Gly, Trp1837Gly, and Trp1837Cys) and 12 in BRCA2 (Leu2510Pro, Asp2611Gly, Tyr2660Asp, Leu2686Pro, Leu2688Pro, Tyr2726Cys, Leu2792Pro, Gly2812Glu, Gly2813Glu, Arg2842Cys, Asp3073Gly, and Gly3076Val) were considered as extremely deleterious. Results suggested that deleterious variants were mostly enriched in theN- and C-terminal domain of the BRCA1 and BRCA2 C-terminus. Utilizing evolutionary conservation analysis, we demonstrated that the majority of deleterious SNPs ensue in highly conserved regions of BRCA genes. Furthermore, utilizing FoldX, we demonstrated that alterations in the function of proteins are not always together with stability alterations.

Methicillin-resistant Staphylococcus aureus (MRSA) is a challenging infectious agent worldwide. The ever growing antibiotic resistance has made the researchers to look for new anti-staphylococcal agents. Autolysins are staphylococcal enzymes that lyse bacterial cell wall for cell division. Autolysins can be used as novel enzybiotics (enzymes have antibiotic effects) for staphylococcal infections. LytU is a newly explored autolysin. SH3b is a potent cell wall binding domain that can be fused to lytic enzymes to increase their activity. The aim of this study was to design a novel and efficient fusion enzybiotic that could lyse staphylococcal cell wall peptidoglycan by disrupting the bacteria. LytU-SH3b fusion construct was synthesized and LytU was amplified through the construct, using overhang PCR. The fusion and native forms that had his-tag were synthesized by recombinant technology in Escherichia coli BL21 (DE3) strain and purified utilizing Ni-NTA agarose beads. LytU and LytU-SH3b activity and potency were assessed using plate lysis assay, turbidity reduction assay and minimal inhibitory concentration (MIC) tests. All these tests showed that LytU-SH3b has more activity and potency than LytU. LytU-SH3b has MIC 421 fold lesser than LytU. Finally, LytU-SH3b is a novel and efficient recombinant enzybiotic that can lyse MRSA as an alternative to chemical small molecule antibiotics.

The prevalence of obesity and diabetes changes dramatically with lifestyle and unequal risk among individuals have made scientists interested to understand how the environment interferes with genetic factors to make it so-called genetic predisposition. This study aimed to explore wherethe most variable region is in leptin gene and analyse  microsatellite repeats with direct sequencing in Iraqis and compare our alleles with other populations as a risk for obesity and T2D predisposition. DNA was extracted from blood of 60 type 2 diabetics and 70 non diabetics individuals, LEP 5‛UTR, exon 2 and 3 were screened in 45 individuals (24 type 2 diabetes patients and 21 non- diabetics), LEP TTTC repeats region were amplified in all 130 participants from which 22 control samples were purified and sequenced, superimposed sequences were analyzed manually. Sequencing results showed G>A polymorphism (rs2167270) in 5‛UTR region. No polymorphisms detected in LEP exons 2 and 3. LEP microsatellites alleles were classified depending on sizes into class1 < (220bp) and class2 (> 220bp). Analysis of 22 control samples sequences of microsatellite region resulted in 6 type1 allele (unique sequence) and 5 type 3 allele (13 different isoforms) depending on TTTC arrangement separated by Ts bases. We concluded that LEP variations were in non- coding regions and no significant difference was observed in allele frequency between both groups, but there was a huge diversity in microsatellite repeat number and context among individuals. This may affects gene function thus prepare a predisposition for obesity and type 2 diabetes.

This research aimed to explore the ACE (insertion/deletion) gene association as key factor for chronic obstructive pulmonary disease (COPD) development in north Indian population. A total of 200 clinically diagnosed patients with COPD were selected against 200 healthy individuals. Genetic variations of ACE (insertion/deletion) were evaluated by using polymerase chain reaction techniques. Smoker showed higher risk of COPD (OR=1.67, 95% CI=1.12-2.48, P=0.012). Present results revealed the positive association between the DD genotype and the risk of COPD (OR= 2.14, 95% CI=1.22-3.78, P=0.006). Among smokers, DD genotype showed statistically significant association with increased risk of COPD (OR=3.10, 95% CI= 1.50-6.47, P=0.001).

Skeletal muscle injuries are one of the most common problems in the worldwide which impose a substantial financial burden to the health care system.  Accordingly, it widely accepted that muscle regeneration is a promising approach that can be used to treat muscle injury patients. However, the underlying mechanisms of muscle regeneration have yet to be elucidated. The muscle structure and muscle-related gene expression are highly conserved between human and zebrafish. Therefore, the zebrafish can be considered as an ideal animal model in muscle regeneration studies. In this study, Tol2 transposase was applied to produce Tg(mylpfa: cfp-nfsB) zebrafish model that express a fusion protein composed of cyan fluorescent protein (CFP) and nitrorudactase (NTR) under control of mylpfa promoter. The results showed that MTZ (Metronidazole) treatment of Tg(mylpfa:cfp-nfsB) zebrafish larvae can lead to muscle injury by selective ablation of muscle cells. And also, results confirmed the muscle regeneration ability of the transgenic larvae after withdrawal of Mtz for three days. Overall, The results of this study suggest that the Tg(mylpfa:cfp-nfsB) zebrafish model can be used in muscle regeneration study in order to elucidate the mechanisms of this process.

Fenugreek is one of the important edible and medicinal vegetables that have a long history of cultivation and consumption. Characterize the extent of the genetic diversity among landraces will provide a good context for future breeding programs and genetic resource preservation. Genetic diversity and population structure of 88 individuals of eight landraces of Iranian fenugreek evaluated based on SRAP markers. Seventy-two bands generated from 6 primers in which 56 (80.11%) band were polymorph. Hamadan landrace showed the lowest values of percentage of polymorphic loci (67.86), Nei's gene diversity index (0.24), number of effective alleles (1.40) and Shannon’s Information index (0.36). Nei’s genetic distance matrix revealed the highest genetic distance between Hamadan and Yazd (0.203) and the highest genetic similarity between Mahallat and Varamin (0.036) landraces. The most gene flow was between Mahallat and Varamin landraces (Nm=8.36) and the least was between Shiraz and Hamadan landraces (Nm=0.66). An extent admixture of alleles between the Iranian fenugreek landraces was observed by the population structure. Mantel test indicated that the genetic differentiation and gene flow is not associated with geographic distance in Iranian fenugreek landraces. Our observations indicated SRAP is an efficient technique to reveal genetic diversity and population structure of Iranian fenugreek landrace.