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Computational insights into fluconazole resistance by the suspected mutations in lanosterol 14α-demethylase (Erg11p) of Candida albicans. 白色念珠菌羊毛甾醇14α-去甲基化酶(Erg11p)可疑突变对氟康唑耐药性的计算见解
IF 1.6 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2020-12-01 DOI: 10.22099/mbrc.2020.36298.1476
Sagunthala Murugesan Udaya Prakash, Yasin Nazeer, Sivaraman Jayanthi, Mohammad Anaul Kabir

Mutations in the ergosterol biosynthesis gene 11 (ERG11) of Candida albicans have been frequently reported in fluconazole-resistant clinical isolates. Exploring the mutations and their effect could provide new insights into the underlying mechanism of fluconazole resistance. Erg11p_Threonine285Alanine (Erg11p_THR285ALA), Erg11p_Leucine321Phenylalanine (Erg11p_LEU321PHE) and Erg11p_Serine457Proline (Erg11p_SER457PRO) are three fluconazole-resistant suspected mutations reported in clinical isolates of C. albicans. Therefore, our study aims to investigate the role of these suspected mutations in fluconazole resistance using in-silico methods. Molecular dynamics simulation (MDS) analysis of apo-protein for 25ns (nanosecond) showed that suspected mutant proteins underwent slight conformational changes in the tertiary structure. Molecular docking with fluconazole followed by binding free energy analysis showed reduced non-bonded interactions with loss of heme interaction and the least binding affinity for Erg11p_SER457PRO mutation. MDS of suspected mutant proteins-fluconazole complexes for 50ns revealed that Erg11p_SER457PRO and Erg11p_LEU321PHE have clear differences in the interaction pattern and loss or reduced heme interaction compared to wild type Erg11p-fluconazole complex. MDS and binding free energy analysis of Erg11p_SER457PRO-fluconazole complex showed the least binding similar to verified mutation Erg11p_TYR447HIS-fluconazole complex. Taken together, our study concludes that suspected mutation Erg11p_THR285ALA may not have any role whereas Erg11p_LEU321PHE could have a moderate role. However, Erg11p_SER457PRO mutation has a strong possibility to play an active role in fluconazole resistance of C. albicans.

白色念珠菌麦角甾醇生物合成基因11 (ERG11)的突变在氟康唑耐药的临床分离株中经常被报道。探索这些突变及其影响可以为氟康唑耐药的潜在机制提供新的见解。Erg11p_Threonine285Alanine (Erg11p_THR285ALA), Erg11p_Leucine321Phenylalanine (Erg11p_LEU321PHE)和Erg11p_Serine457Proline (Erg11p_SER457PRO)是在临床分离的白色念菌中报道的三种氟唑耐药可疑突变。因此,我们的研究旨在利用计算机方法研究这些可疑突变在氟康唑耐药中的作用。载脂蛋白25ns(纳秒)的分子动力学模拟(MDS)分析表明,疑似突变蛋白在三级结构上发生了轻微的构象变化。结合自由能分析显示,与氟康唑的分子对接减少了非键相互作用,失去了血红素相互作用,对Erg11p_SER457PRO突变的结合亲和力最低。疑似突变蛋白-氟康唑复合物50ns的MDS结果显示,与野生型erg11p -氟康唑复合物相比,Erg11p_SER457PRO和Erg11p_LEU321PHE在相互作用模式和血红素相互作用缺失或减少方面存在明显差异。MDS和结合自由能分析显示erg11p_ser457pro -氟康唑复合物的结合最少,与已验证的突变erg11p_tyr447his -氟康唑复合物相似。综上所述,我们的研究得出结论,疑似突变Erg11p_THR285ALA可能没有任何作用,而Erg11p_LEU321PHE可能有中等作用。而Erg11p_SER457PRO突变极有可能在白念珠菌氟康唑耐药中发挥积极作用。
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引用次数: 3
Differential expression of inflammatory responsive genes between chronic periodontitis and periodontally affected bronchiectasis patients. 慢性牙周炎与牙周影响支气管扩张患者炎症反应基因的差异表达。
IF 1.6 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2020-12-01 DOI: 10.22099/mbrc.2020.37397.1508
Abhaya Gupta, Neetu Singh, Anil Kumar, Umesh Pratap Verma, Ajay Kumar Verma, Hari Shyam, Nand Lal, Surya Kant, Ankur Kumari

The study aimed to investigate differential expression of targeted inflammatory-immune responsive genes [LTA, LTB, TNFSF4, TNFSF11/RANKL, TNFSF13, TNFSF13B, TNFRSF11B/ Osteoprotegerin; OPG and GFPT1/GFA ] in gingival tissues of bronchiectasis patients having chronic periodontitis in North central Indian population. Gingival tissues were collected from 30 systemically healthy chronic periodontitis patients (CP), 30 bronchiectasis patients with chronic periodontitis (B+CP), 3 systemically healthy with healthy gingiva (healthy control; HC) and 3 bronchiectasis with healthy gingiva (bronchiectasis control; BC). Statistical analysis revealed 7 genes to be significantly upregulated on comparing CP with B+CP i.e LTA (P<0.0001) in B+CP while LTB (P<0.0001), TNFSF4 (P=0.0003), TNFSF11 (P<0.0001), TNFSF13 (P=0.0003), TNFSF13B (P<0.0001) and TNFRSF11B (P=0.0004) in CP group. LTA (Lymphotoxin A) gene could be a potential genetic marker in bronchiectasis patients with chronic periodontitis.

本研究旨在探讨靶向炎症免疫应答基因[LTA、LTB、TNFSF4、TNFSF11/RANKL、TNFSF13、TNFSF13B、TNFRSF11B/ Osteoprotegerin;印度中北部人群慢性牙周炎支气管扩张患者牙龈组织的OPG和GFPT1/GFA。收集30例全身健康的慢性牙周炎患者(CP)、30例支气管扩张合并慢性牙周炎(B+CP)、3例全身健康且牙龈健康的患者(健康对照组;HC)和3例健康牙龈的支气管扩张(支气管扩张对照组;BC)。经统计学分析,CP组与B+CP组比较,有7个基因显著上调,分别为LTA (PLTB) (PTNFSF4) (P=0.0003)、TNFSF11 (PTNFSF13) (P=0.0003)、TNFSF13B (PTNFRSF11B) (P=0.0004)。LTA(淋巴毒素A)基因可能是支气管扩张合并慢性牙周炎的潜在遗传标记。
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引用次数: 1
Differential genes expression analysis of invasive aspergillosis: a bioinformatics study based on mRNA/microRNA. 侵袭性曲霉病差异基因表达分析:基于mRNA/microRNA的生物信息学研究。
IF 1.6 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2020-12-01 DOI: 10.22099/mbrc.2020.37432.1509
Maryam Hosseinipour, Shirin Shahbazi, Shahla Roudbar-Mohammadi, Maryam Khorasani, Majid Marjani

Invasive aspergillosis is a severe opportunistic infection with high mortality in immunocompromised patients. Recently, the roles of microRNAs have been taken into consideration in the immune system and inflammatory responses. Using bioinformatics approaches, we aimed to study the microRNAs related to invasive aspergillosis to understand the molecular pathways involved in the disease pathogenesis. Data were extracted from the gene expression omnibus (GEO) database. We proposed 3 differentially expressed genes; S100B, TDRD9 and TMTC1 related to pathogenesis of invasive aspergillosis. Using miRWalk 2.0 predictive tool, microRNAs that targeted the selected genes were identified. The roles of microRNAs were investigated by microRNA target prediction and molecular pathways analysis. The significance of combined expression changes in selected genes was analyzed by ROC curves study. Thirty-three microRNAs were identified as the common regulator of S100B, TDRD9 and TMTC1 genes. Several of them were previously reported in the pathogenesis of fungal infections including miR-132. Predicted microRNAs were involved in innate immune response as well as toll-like receptor signaling. Most of the microRNAs were also linked to platelet activation. The ROC chart in the combination mode of S100B/TMTC1, showed the sensitivity of 95.65 percent and the specificity of 69.23 percent. New approaches are needed for rapid and accurate detection of invasive aspergillosis. Given the pivotal signaling pathways involved, predicted microRNAs can be considered as the potential candidates of the disease diagnosis. Further investigation of the microRNAs expression changes and related pathways would lead to identifying the effective biomarkers for IA detection.

侵袭性曲霉病是一种严重的机会性感染,免疫功能低下患者死亡率高。近年来,microrna在免疫系统和炎症反应中的作用已被考虑。利用生物信息学方法,我们旨在研究与侵袭性曲霉病相关的microrna,以了解疾病发病机制中涉及的分子途径。数据从基因表达综合数据库(GEO)中提取。我们提出了3个差异表达基因;S100B、TDRD9和TMTC1与侵袭性曲霉病发病有关。使用miRWalk 2.0预测工具,鉴定了靶向所选基因的microrna。通过microRNA靶点预测和分子通路分析研究了microRNA的作用。采用ROC曲线分析所选基因联合表达变化的意义。33个microrna被鉴定为S100B、TDRD9和TMTC1基因的共同调控因子。其中一些先前在真菌感染的发病机制中被报道过,包括miR-132。预测的microrna参与先天免疫反应以及toll样受体信号传导。大多数microrna也与血小板活化有关。S100B/TMTC1联合模式下的ROC图显示灵敏度为95.65%,特异度为69.23%。需要新的方法来快速准确地检测侵袭性曲霉病。考虑到所涉及的关键信号通路,预测的microrna可以被认为是疾病诊断的潜在候选者。进一步研究microrna的表达变化及其相关途径将有助于确定IA检测的有效生物标志物。
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引用次数: 1
Association of two single nucleotide polymorphisms rs10407022 and rs3741664 with the risk of primary ovarian insufficiency in a sample of Iraqi women. 伊拉克妇女样本中两个单核苷酸多态性rs10407022和rs3741664与原发性卵巢功能不全风险的关联
IF 1.6 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2020-12-01 DOI: 10.22099/mbrc.2020.36371.1477
Tamadher Abbas Rafaa, Ahmed AbdulJabbar Suleiman, Mustafa Falah Dawood, Ali Mohammed Al-Rawi

Primary ovarian insufficiency (POI) can be a devastating disease impacting women below the age of forty. This involves a major decrease in the amount and quality of oocytes, or ovarian reserve in a woman. The distribution of single-nucleotide polymorphisms, rs10407022 and rs3741664, in Iraqi people and its association with primary ovarian insufficiency is the main objective of this study. The mean of FSH and LH levels of patients with POI was higher than control, while the mean of AMH levels of patients was lower compared to control. For rs10407022, the GT and TT genotypes were positively associated with the risk of POI. For the rs3741664, the AG genotype was negatively associated with the risk of POI. The results lead to the main conclusion that rs10407022 and rs3741664 polymorphisms may significantly affected the serum levels of AMH and FSH and thus affect POI etiology.

原发性卵巢功能不全(POI)是一种影响40岁以下女性的毁灭性疾病。这涉及到女性的卵母细胞或卵巢储备的数量和质量的显著下降。单核苷酸多态性rs10407022和rs3741664在伊拉克人中的分布及其与原发性卵巢功能不全的关系是本研究的主要目的。POI患者FSH、LH水平均值高于对照组,AMH水平均值低于对照组。对于rs10407022, GT和TT基因型与POI风险呈正相关。对于rs3741664, AG基因型与POI风险呈负相关。结果表明rs10407022和rs3741664多态性可能显著影响血清AMH和FSH水平,从而影响POI病因。
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引用次数: 0
Prevalence of Brucella species in unpasteurized dairy products consumed in Shiraz province using PCR assay. 用PCR法测定设拉子省消费的未经巴氏消毒的乳制品中布鲁氏菌的流行情况。
IF 1.6 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2020-09-01 DOI: 10.22099/mbrc.2020.37381.1506
Fargol Abdali, Saeid Hosseinzadeh, Enayat Berizi, Maryam Pourmontaseri

The consumption of milk and unpasteurized dairy products contaminated with Brucella bacteria is one of the most important ways of brucellosis transmission to humans. The principal goal of this study was to determine the prevalence of Brucella abortus (B. abortus) and Brucella melitens (B. melitens) in unpasteurized dairy products consumed in Shiraz province. In this study conducted in 2016, 238 unpasteurized dairy products including 48 raw milk, 48 yogurt, 46 cheeses, 48 dough and 48 ice cream samples, were purchased from the retail market in Shiraz province and were examined by a specific PCR assay. This study showed positive 5/04% out of 238 unpasteurized dairy products including 9 out of 48 (18/75%) raw milk samples and 3 out of 48 (6.25%) yogurt samples). Contamination was not detected in samples of dough, cheese and traditional ice cream. The results also showed that among 12 positive samples, 6 samples were contaminated with B. abortus (including 4 milk samples and 2 yogurt samples), 2 samples were contaminated with B. melitensis (including 2 Milk samples) and 4 samples were contaminated simultaneously with B. abortus and B. melitensis (including 3 milk samples and 1 yogurt sample). The present study suggests the unpasteurized dairy products as the major sources of brucellosis in Shiraz province, South of Iran; thus, to prevent brucellosis in human, the consumption of pasteurized milk and dairy products is highly recommended.

食用被布鲁氏菌污染的牛奶和未经巴氏消毒的乳制品是布鲁氏菌病传播给人类的最重要途径之一。本研究的主要目的是确定在设拉子省消费的未经高温消毒的乳制品中流产布鲁氏菌(B. abortus)和melitens布鲁氏菌(B. melitens)的患病率。在2016年进行的这项研究中,从设拉子省的零售市场购买了238种未经巴氏消毒的乳制品,包括48种生乳、48种酸奶、46种奶酪、48种面团和48种冰淇淋样品,并通过特定的PCR检测进行了检测。该研究显示,238种未经巴氏消毒的乳制品中有5/04%呈阳性,其中包括48种原料奶样品中的9种(18/75%)和48种酸奶样品中的3种(6.25%)。在面团、奶酪和传统冰淇淋的样本中没有检测到污染。结果还显示,在12份阳性样品中,6份样品被产弧菌污染(包括4份牛奶样品和2份酸奶样品),2份样品被melitensis污染(包括2份牛奶样品),4份样品同时被产弧菌和melitensis污染(包括3份牛奶样品和1份酸奶样品)。目前的研究表明,未经高温消毒的乳制品是伊朗南部设拉子省布鲁氏菌病的主要来源;因此,为预防人类感染布鲁氏菌病,强烈建议食用巴氏奶和奶制品。
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引用次数: 3
Sequence variants  of  CYP345a1 and CYP6a14 gene regions in Tribolium castaneum (Coleoptera: Tenebrionidae) adults treated with the novel characterızed Bolanthus turcicus (Caryophyllaceae) extract. 新提取物characterızed处理后的黄褐虫(鞘翅目:拟甲科)成虫CYP345a1和CYP6a14基因区序列变异
IF 1.6 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2020-09-01 DOI: 10.22099/mbrc.2020.35861.1472
Fahriye Sümer Ercan, Serap Yalçın Azarkan, Nuri Ercan, Murat Koc

In this study, various doses of plant extracts that obtained from Bolanthus turcicus was applied to an important storage pest Tribolium castaneum adults. Bolanthus turcicus is an endemic species and spreads on the Hasan Mountain above Karkın town (Turkey, Aksaray province). The plant species was collected from June to July with the field study to be carried out in this region. Obtained extract of plant was analyzed by gas chromatography mass spectrometry (GC-MS) method. The doses were defined during the study and the concentrations that kill 50% and 99% of the population were determined after applications. After 24 h, DNA was isolated from live and dead individuals that obtained from LC50 and LC99 concentration applications and analyzed for Cytochrome P450-mediated detoxification resistance genes, CYP345A1 and CYP6A14 gene regions, by polymerase chain reaction (PCR). CYP genes in insects are known to be rapidly regulated when exposed to insecticides. In the study, in order to screen for 206 bp and 353 bp fragments of CYP345A1 and CYP6A14 genes in T. castaneum adults were amplified using specific primers, respectively. DNA direct sequencing was performed on each template using the forward primer. When compared to the control, it is believed that mutation differences in live and dead individuals according to the sequencing results obtained from survival and dead adults, may allow these genes to play a protective role against the toxic effect of B. turcicus extract.

本研究采用不同剂量的黄菖蒲提取物对重要的储存性害虫黄菖蒲成虫进行了防治。Bolanthus turcicus是一种特有种,分布在Karkın镇(土耳其,Aksaray省)上方的Hasan山上。植物种类采集于6 - 7月,野外研究将在该地区进行。采用气相色谱-质谱(GC-MS)法对提取的植物提取物进行分析。在研究期间确定了剂量,并在施用后确定了杀死50%和99%人口的浓度。24 h后,从LC50和LC99浓度应用中获得的活个体和死个体中分离DNA,采用聚合酶链反应(PCR)分析细胞色素p450介导的解毒抗性基因CYP345A1和CYP6A14基因区域。昆虫体内的CYP基因在暴露于杀虫剂时被迅速调节。本研究利用特异性引物分别扩增了castaneum成虫CYP345A1和CYP6A14基因的206 bp和353 bp片段,以筛选其基因序列。使用正向引物对每个模板进行DNA直接测序。与对照相比,根据从存活和死亡的成虫中获得的测序结果,认为活的和死的个体的突变差异可能使这些基因对黄芽孢杆菌提取物的毒性作用发挥保护作用。
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引用次数: 3
Deciphering the functional role of hypothetical proteins from Chloroflexus aurantiacs J-10-f1 using bioinformatics approach. 用生物信息学方法解读金莲J-10-f1假想蛋白的功能作用。
IF 1.6 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2020-09-01 DOI: 10.22099/mbrc.2020.36894.1495
Chander Jyoti Thakur, Sandeep Saini, Aayushi Notra, Bhavanshu Chauhan, Sarthak Arya, Rishabh Gupta, Jyotsna Thakur, Varinder Kumar

Chloroflexus aurantiacus J-10-f1 is an anoxygenic, photosynthetic, facultative autotrophic gram negative bacterium found from hot spring at a temperature range of 50-60°C. It can sustain itself in dark only if oxygen is available thereby exhibiting a dark orange color, however display a dark green color when grown in sunlight. Genome of the organism contains total of 3853 proteins out of which 785 (~20%) proteins are uncharacterised or hypothetical proteins (HPs). Therefore in this work we have characterized the 785 hypothetical proteins of Chloroflexus aurantiacus J-10-f1 using bioinformatics tools and databases. HPs annotated by more than five domain prediction tools were filtered and named high confidence-hypothetical proteins (HC-HPs). These HC-HPs were further annotated by calculating their physiochemical properties, homologous, subcellular locations, signal peptides and transmembrane regions. We found most of the HC-HPs were involved in photosynthesis, carbohydrate metabolism, biofuel production and cellulose synthesis processes. Furthermore, few of these HC-HPs could provide resistance to bacteria at high temperature due to their thermophilic nature. Hence these HC-HPs have the potential to be used in industrial as well as in biomedical needs. To conclude, the bioinformatics approach used in this study provides an insight to better understand the nature and role of Chloroflexus aurantiacus J-10-f1 hypothetical proteins.

aurantiacus J-10-f1是一种无氧、光合、兼性自养革兰氏阴性细菌,发现于温度范围为50-60℃的温泉中。它只有在有氧气的情况下才能在黑暗中生存,因此呈现出深橙色,然而在阳光下生长时呈现出深绿色。生物基因组共包含3853种蛋白质,其中785种(约20%)蛋白质是未表征或假设的蛋白质(HPs)。因此,本研究利用生物信息学工具和数据库,对黄颡鱼J-10-f1的785个假设蛋白进行了表征。被5个以上结构域预测工具注释的hp被过滤并命名为高置信度假设蛋白(hc - hp)。通过计算其理化性质、同源性、亚细胞位置、信号肽和跨膜区域,进一步对这些hc - hp进行注释。我们发现大多数hc - hp参与光合作用、碳水化合物代谢、生物燃料生产和纤维素合成过程。此外,由于这些hc - hp的嗜热性,它们很少能在高温下对细菌产生抗性。因此,这些hc - hp具有用于工业和生物医学需求的潜力。综上所述,本研究采用的生物信息学方法有助于更好地了解黄绿J-10-f1假设蛋白的性质和作用。
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引用次数: 5
Evaluation of the risk of lung cancer associated with NAD(P)H: quinone oxidoreductase 1 (NQO1) C609T polymorphism in male current cigarette smokers from the Eastern India. 东印度男性吸烟者NAD(P)H:醌氧化还原酶1 (NQO1) C609T多态性与肺癌风险的评估
IF 1.6 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2020-09-01 DOI: 10.22099/mbrc.2020.36467.1481
Santanu Banerjee

NAD(P)H: quinone oxidoreductase 1 (NQO1) is an endogenous cellular defence mechanism against several carcinogenic quinones derived from cigarette smoke. NQO1 C609T polymorphism is a strong determinant of NQO1 structure and function. The people with mutant allele for this polymorphism has significantly reduced NQO1 activity. In this study, we tried to evaluate the risk of lung cancer associated with this polymorphism in male current smokers of the Eastern India. Using PCR-RFLP method, we compared the NQO1 C609T genotype distribution in male current smokers with (n=150) and without (n=200) lung cancer. We observed significant variation of genotypic distribution between these two groups. The allele frequency of the variant C609T allele were 40.3% and 32.7% in smokers with and without lung cancer, respectively. From the genotypic comparison between the two smoker groups, it was found that a higher risk (OR=1.64, 95% CI: 1.05-2.55, P<0.05) of lung cancer was associated with NQO1 C609T polymorphism.

NAD(P)H:醌氧化还原酶1 (NQO1)是一种内源性细胞防御机制,可抵抗来自香烟烟雾的几种致癌醌。NQO1 C609T多态性是NQO1结构和功能的重要决定因素。携带该多态性等位基因突变的人,其NQO1活性显著降低。在这项研究中,我们试图评估与这种多态性相关的肺癌风险在印度东部的男性吸烟者中。采用PCR-RFLP方法,比较了肺癌男性吸烟者(n=150)和非肺癌男性吸烟者(n=200) NQO1 C609T基因型分布。我们观察到两组间基因型分布有显著差异。C609T等位基因变异频率在肺癌吸烟者和非肺癌吸烟者中分别为40.3%和32.7%。从两组吸烟者的基因型比较中,发现PNQO1 C609T多态性的风险更高(OR=1.64, 95% CI: 1.05-2.55)。
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引用次数: 0
A novel chimeric recombinant protein PDHB-P80 of Mycoplasma agalactiae as a potential diagnostic tool. 一种新的无乳支原体嵌合重组蛋白PDHB-P80作为潜在的诊断工具。
IF 1.6 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2020-09-01 DOI: 10.22099/mbrc.2020.37684.1513
Malihe Akbarzadeh-Niaki, Abdollah Derakhshandeh, Nasrin Kazemipour, Vida Eraghi, Farhid Hemmatzadeh

The aim of this study was to construct, expression of a novel recombinant chimeric protein consisting of Pyruvate dehydrogenase beta subunit (PDHB) and high antigenic region of integral membrane lipoprotein P80 of Mycoplasma agalactiae as a potential diagnostic tool. The full-length sequence of pdhb and a portion of antigenic regions of P80 were selected and analyzed by CLC main workbench 5.5 software. Several linkers and three dimensional structure of PDHB-P80 were compared to the native PDHB and analyzed to select a proper one for expression. The fusion gene sequence was optimized and synthesized in pMAT cloning vector. The synthetic pMAT-pdhb-p80 was digested using Bam HI and Sal I restriction enzymes and ligated into pMAL-p5X expression vector. The pMAL-pdhb-p80 construct was transfected into E.coli BL21 strain cells and expressed protein were purified using amylose resin. and the purified protein was analyzed in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting. In silico analysis demonstrated that fusion proteins using IgG4 middle hinge (CPSCP) with TM-score of 0.99 showed the higher similarity between three dimensional structure of PDHB before and after fusion with high antigenic region of P80. Successful cloning verified by PCR colony, double digestion and sequence analysis. Besides, SDS-PAGE analysis and Western blotting indicated and confirmed the expression of intact recombinant chimeric protein MBP-PDHB-P80 along with some truncated forms of the recombinant protein. it could be concluded that the fusion construct has a potential for serodiagnostic assay in future studies.

本研究的目的是构建和表达一种由丙酮酸脱氢酶β亚基(PDHB)和无乳支原体整体膜脂蛋白P80高抗原区组成的新型重组嵌合蛋白,作为一种潜在的诊断工具。选取pdhb的全长序列和P80的部分抗原区,利用CLC主工作台5.5软件进行分析。将PDHB- p80的几种连接体和三维结构与天然PDHB进行比较,并分析选择合适的表达载体。在pMAT克隆载体上优化并合成了融合基因序列。合成的pMAT-pdhb-p80用Bam HI和Sal I酶切后连接到pmat - p5x表达载体上。将pMAL-pdhb-p80构建体转染大肠杆菌BL21细胞,用直链淀粉树脂纯化表达蛋白。用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)和Western blotting对纯化蛋白进行分析。计算机分析表明,使用tm评分为0.99的IgG4中间枢纽(CPSCP)融合蛋白与P80高抗原区融合前后,PDHB的三维结构具有较高的相似性。经PCR菌落、双酶切和序列分析证实克隆成功。此外,SDS-PAGE分析和Western blotting显示并证实了完整的重组嵌合蛋白MBP-PDHB-P80的表达以及重组蛋白的一些截短形式。由此可见,该融合构建体在今后的研究中具有用于血清诊断的潜力。
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引用次数: 2
Using two retrotransposon-based marker systems (SRAP and REMAP) for genetic diversity analysis of Moroccan Argan tree. 利用两个基于反转录转座子的标记系统(SRAP和REMAP)对摩洛哥坚果树进行遗传多样性分析。
IF 1.6 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2020-09-01 DOI: 10.22099/mbrc.2020.36390.1478
Ouafae Pakhrou, Leila Medraoui, Bouchra Belkadi, Farid Rachidi, Hasnaa Errahmani, Mohammed Alami, Abdelkarim Filali-Maltouf

The Argania is an endemic genetic resource in Morocco holding an important ecological and socio-economical benefit. However, overgrazing and overharvesting lead to a serious downturn in the number of trees. To characterize genetic diversity within and among 24 populations, represented by 240 argan trees, four combinations of SRAP primers and eight combinations of REMAP primers were used. A total of 338 REMAP and 146 SRAP markers were amplified with a polymorphism of 100%. The average polymorphism information content value was 0.20 and 0.17 for SRAP and REMAP markers, respectively. The analysis of molecular variance showed that 26% of the genetic variation was partitioned among populations. The coefficient of gene differentiation was 0.2875 and gene flow was 1.2391. The average parameter diversity was: observed number of alleles (Na)=0.729, effective number of alleles (Ne)=1.131, Shannon's information index (I)=1.143; Nei's gene diversity (H)=0.093 and Percentage of Polymorphic Loci=35.68. The STRUCTURE and principal coordinate analysis revealed that the Argania spinosa L. populations were aggregated into 2 genetic groups. To detect outlier, baysecan software was used and 21 were detected (7 under selection, 14 under balancing selection) presenting posterior probability higher than 0.79. The current results can be explored in the design of management programs and to comprehend the adaptation mechanism of Argan tree.

阿根尼亚是摩洛哥特有的遗传资源,具有重要的生态效益和社会经济效益。然而,过度放牧和过度采伐导致树木数量严重下降。为了表征240棵摩洛哥坚果树所代表的24个群体内部和群体间的遗传多样性,使用了4种SRAP引物组合和8种REMAP引物组合。共有338个REMAP标记和146个SRAP标记扩增,多态性为100%。SRAP和REMAP标记的平均多态性信息含量值分别为0.20和0.17。分子方差分析表明,26%的遗传变异在群体间被划分。基因分化系数为0.2875,基因流量为1.2391。平均参数多样性为:观察等位基因数(Na)=0.729,有效等位基因数(Ne)=1.131,香农信息指数(I)=1.143;Nei’s基因多样性(H)=0.093,多态性位点百分率=35.68。结构分析和主坐标分析表明,棘蚶居群可聚集为2个遗传群。使用baysecan软件检测异常值,检测到21例(选择7例,平衡选择14例),后验概率大于0.79。目前的研究结果可为管理方案的设计和了解摩洛哥坚果树的适应机制提供参考。
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引用次数: 6
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Molecular Biology Research Communications
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