生物活性有机锗及HPCPC双水相分离生物大分子的研究

本论文内容主要分为4个部分：“有机锗化合物抑制Maillard反应的研究”、“Ge-132对体外培养幼鼠胰岛细胞的作用研究”、“PEG／磷酸盐双水相体系中生物大分子分配的研究”和“HPCPC（High Performance Centrifugal Partition Chromatography，高效离心分配色谱）双水相体系对生物大分子分离的研究”。前两部分主要从有机锗化合物防治糖尿病及其并发症的代谢机理和细胞学角度进行研究，本文引入HPCPC和双水相体系分离生物大分子的技术，为进一步阐明有机锗化合物在Maillard反应过程中的作用机理：对双水相体系中生物大分子的分配及HPCPC在双水相体系中分离生物大分子也做了大量的基础性研究，为开发HPCPC的应用作出了有价值的探索工作。本论文的主要创新点归纳如下：一、有机锗化合物对Maillard反应的抑制作用：针对国际上有机锗发展的前沿课题，在国内首先开展了有机锗化合物对Maillard反应抑制作用的研究，取得了有价值的研究结果。1、在模拟体内的生理条件下，研究了不同浓度Ge-132对精氨酸、组氨酸、甘氨酸和缬氨酸对Maillard反应荧光峰强度的影响：不同类型的氨基酸Maillard反应产物结构上的差异和不同浓度有机锗对组氨酸糖化反应420nm处特征吸光度的影响。2、苯丙氨酸是侧链带有苯环的芳香族氨基酸，是一种具有弱的天然荧光的生物小分子，其荧光最大发射位置在281.6nm。由于aillard反应是葡萄糖和氨基酸的-NHR基发生的反应，其反应产物的特征荧光在440nm附近。这两类荧光的发射位置相差较远，相互之间没有影响，因此利用荧光法观察Phe在反应过程中自身的荧光变化和Ge-132对Phe的Maillard反应荧光产物的抑制情况。3、血清白蛋白是哺乳动物体内的重要蛋白质，可作为多种内源性、外源性物质的存储和转运蛋白，其Maillard反应已经被广泛重视，但较复杂的实验条件限制了它的研究。通过观测蛋白质Maillard反应产物特征荧光的变化是一有效的研究方法。BSA的内源性荧光是由肽链色氨酸和酪氨酸残基贡献的，其最大发射位置340nm左右。其Maillard反应产物的特征荧光在440nm附近，这两类荧光发射位置相差较远，基本上没有影响，因此能够通过荧光光谱研究BSA的Maillard反应荧光产物的情况。本文观测了Ge-132对BSA的Maillard反应荧光产物的抑制作用，同时还通过荧光法确定了一个文献中没有的新的反应位点。4、本文研究了具有更高水溶性的一类双有机锗化合物(HO)_2Ge(CHR_1CHR_2COOH)_2．2H_20抑制Maillard反应的特性，结果证明它比Ge-132具有更高的抑制作用，我们通过化合物的构效关系解释了上述作用的机制。5、本文研究了Tb(Ge-132)_3和Eu(Ge-132)_3两种含稀土的配合物对氨基酸、蛋白质Maillard反应的影响，观察了稀土离子对Maillard反应体系的影响，对实验现象作出了合理的解释。二、Ge-132对体外培养幼鼠胰岛细胞的作用为进一步考察有机锗对糖尿病的防治作用，我们与白求恩医大合作，首次观察了Ge-132对体外培养的幼鼠胰岛细胞结构和功能的影响，研究了不同剂量的含锗化合物对幼鼠胰岛细胞分泌胰岛素的作用，认为低浓度的Ge-132对胰岛细胞分泌胰岛素具有明显的促进作用。该部分内容为首次报导。三、PEG／磷酸盐双水相体系中生物大分子分配的研究双水相萃取TPE(Aqueous Two-Phase Extraction)具有下述优点：生物相容性好、界面张力低、能量低、易于工业规模的放大、可以进行连续操作等。本论文系统地研究了各种蛋白质在PEG／磷酸盐体系中的分离情况，如在各种pH条件下，从6.8到9.2，各种分子量的PEG，包括PEGl000，2000，5000，6000，10000，和20000和磷酸盐双水相体系的物化性质，以及对lysozyme（溶菌酶），BSA（牛血清白蛋白），HSA（人血清白蛋白），Hemoglobin（血红蛋白）在各种体系中的分配系数，探讨了各种因素对生物大分子分配的影响。四、HPCPC在双水相体系对生物大分子分离的研究由于荧光光谱法证实了Ge-132对牛血清白蛋白Maillard反应的抑制作用。但对于如何能够分离出糖化产物，达到进行定量表征的目的是我们研究的一个重要内容。九十年代初，由日本研制出一种色谱中新技术HPCPC。HPCPC与传统的液相色谱(LC)和高效液相色谱(HPLC)不同，不需要固体作为支持体。流动相和固定相分别为两种不相混溶的液体，通过离心力的作用使其中一相作为固定相保持在类似多级萃取器的微小分配槽中，另一相作为流动相流经固定相。HPCPC在双水相体系的应用是一种利用多级连续萃取从双水相中分离生物物质的方法它具有运行时间快、高效高选择性，流动相与固定相之间比例可以任意改变，并在任何pH值均可进行正向和反向操作并且实验室规模可一步直接放大到生产规模。它不仅可以作为一种分离的工具，而且也是一种研究生物萃取反应动力学机理，反映热力学与动力学之间相关性的重要方法。本论文中研究了常用的两种双水相体系，利用PEG6000／Dex20000／H_20对BSA和Maillard反应产生的糖化BSA进行了初步分离，取得了一些阶段性结论。另外，通过对各种条件的筛选，选择了具有代表性的lysozyme和BSA，BSA和血红蛋白(Hb)进行HPCPC色谱仪上的分离，研究了色谱分离条件如：转数、流速、pH等对分离效率及理论塔板数的影响，取得了一系列重要的结论。为工业上大规模的生物样品分离提供了重要的参数。

毛壳菌产抗真菌活性物质菌株筛选及种间原生质体融合研究

毛壳菌属很多种类具有重要生防价值，其生防机理包括对植物病原真菌的重寄生作用、诱导植物产生抗病性、产生抗真菌活性的次生代谢产物等。迄今，学界对毛壳菌的研究主要集中在毛壳菌的生防机理，毛壳菌活性次生代谢产物的分离等方面。本研究致力于产抗生素的毛壳菌的种间原生质体融合，从产抗生素毛壳菌菌株的筛选开始，进而对产抗生素的角毛壳菌进行诱变选育，最终用产不同抗生素的角毛壳菌与球毛壳菌进行种间原生质体融合。主要有以下五方面研究结果。 1、毛壳菌抗真菌活性物质产生菌株的筛选：不同毛壳菌菌株发酵液采用琼脂扩散法对植物病原真菌进行抑菌活性试验，结果显示，菌株CH08和CH23的发酵液对芒果炭疽、苹果炭疽和马铃薯晚疫菌具有抑制作用。菌株CH16和CH17的发酵液对芒果炭疽菌、苹果炭疽菌有抑制作用。菌株CH21发酵液对辣椒炭疽菌和西瓜枯萎菌有抑制作用。经形态学研究，菌株CH08、CH16、CH17和CH23鉴定为球毛壳菌，菌株CH21鉴定为角毛壳菌。对角毛壳菌与球毛壳菌菌株发酵液抑菌谱比较，发现角毛壳菌与球毛壳菌发酵液具有明显不同的抑菌谱，表明角毛壳菌与球毛壳菌产生不同的抗真菌活性物质。 2、角毛壳菌（CH21）和球毛壳菌（CH08）原生质体制备和再生条件研究：考察了菌龄、酶浓度、稳渗剂及其浓度、酶解温度、酶解时间及再生培养基对原生质体制备和再生的影响。用菌龄为生长54 h的角毛壳菌菌丝，以0.06 M磷酸缓冲液（pH6.0）配制成含蜗牛酶15 mg/ml、溶壁酶10 mg/ml、蔗糖0.6 mol/L的酶解液，30℃酶解1.5 h，原生质体释放量2.02×107个/g；以PDA为再生培养基，0.7 mol/L的蔗糖再生稳渗剂，再生率可达51.45%。用菌龄为生长48 h的球毛壳菌菌丝，以0.06 M磷酸缓冲液（pH6.0）配制成含蜗牛酶15 mg/ml、溶壁酶10 mg/ml、蔗糖0.6 mol/L的酶解液，30℃酶解1 h，原生质体释放量达1.57×108个/g；以PDA为再生培养基，0.7 mol/L的蔗糖为再生稳渗剂，再生率可达41.48%。 3、角毛壳菌（CH21）原生质体紫外诱变选育：以CH21为出发菌株，制备原生质体进行紫外诱变，诱变条件为：15 w紫外灯，距离30 cm，照射90 s，致死率80%~85%。建立了诱变菌株初筛的双层平板筛选模型。经平板初筛和摇瓶复筛，获得一株突变菌株CH21-I-402，其发酵液抑菌活性较出发菌株提高18.3%。 4、抗性标记菌株的获得：菌株CH21-I-402和CH08抗生素药敏试验表明， CH21-I-402菌株对潮霉素有抗性、对G418（Geneticin）敏感，菌株CH08对潮霉素和G418都敏感。根癌农杆菌EHA105介导的新霉素磷酸转移酶基因转化球毛壳菌，经PCR检测，新霉素磷酸转移酶基因成功转化进菌株CH08-GR70，CH08-GR120。转化子对G418抗性提高3～4倍，对潮霉素仍然比较敏感。 5、以G418和潮霉素抗性为筛选标记的原生质体融合与融合菌株AFLP分析：制备角毛壳菌CH21-I-402和球毛壳菌CH08-GR70原生质体，以35％的PEG6000为助融剂进行原生质体融合，以65 μg/ml的潮霉素和60 μg/ml G418为抗性筛选标记，获得46个再生菌株。再生菌株连续传代5代后，再生菌株表现出多种形态类型。利用AFLP技术对再生菌株及亲本菌株基因组DNA分析表明，再生菌株PF1、PF26为融合菌株。抑菌活性测试表明，融合菌株PF26发酵液对芒果炭疽菌和苹果轮纹菌有强的抑制作用，且抑菌活性比亲本球毛壳菌明显提高。 Chaetomium spp. have great potentials as biocontrol agents against a range of plant pathogens on the basis of its mycoparasitism, induced plant disease resistance, production of antifungal metabolites, and so on. Previous researches on C. spp. mostly focused on the mechanisms of its biocontrol and the isolation of secondary metabolites. In this study, screening antifungal C. spp., mutation breeding of C. cupreum and interspecies protoplast fusion between C. cupreum and C. globosum were carried out, respectively. The corresponding results are as follows: Firstly, among more than 40 C. spp., the strains produced anti-fungal antibiotics were screened by agar diffusion experiments. Results showed that both CH08 and CH23 had inhibition against Colletotrichum gloeosporioides, Cladosporium fulvum, and Phytophthora infestans. Both CH16 and CH17 had inhibition against Colletotrichum gloeosporioides and Cladosporium fulvum. In addition, CH21 exhibited anti-fungal activity against Fusarium oxysporum f. sp niveum and Colletotrichum capsici. Furthermore, CH08, CH16, CH17 and CH23 were identified as C. globosum, CH21 was proved to be C. cupreum based on morphology. The comparison of the anti-fungal spectrum between C. cupreum and C. globosum, showed they could produce different antibiotics. Secondly, specified protocols for preparing and regenerating protoplasts from mycelia of C. cupreum CH21 and C. globosum CH08 were studied. The effects of the age mycelia, the concentration of enzyme, digestion temperature and time, kinds of osmotic stabilizer and regeneration medium on protoplasts preparation and regeneration were all optimized, respectively. In one protocol, with 15 mg/mL snailase, 10 mg/mL lywallzyme, 0.6 M sucrose, in 0.06 M phosphate buffer (pH6.0), and digested for 1.5 h at 30 ºC, 2.02×107 protoplasts from each gram mycelia were obtained from cultures of C. cupreum CH21 grown in potato dextrose broth (PDB) medium for 54 h. And when 0.7 M sucrose was used as osmotic stabilizer in the regeneration medium OPDA (potato dextrose agar with osmotic stabilize), the regeneration efficiency of protoplasts was 51.45%. In another protocol, with 15 mg/mL snailase, 10 mg/mL lywallzyme, 0.6 M sucrose, in 0.06 M phosphate buffer (pH6.0), and digested for 1 h at 30 ºC, 1.57×108 protoplasts from each gram mycelia were obtained from cultures of C. globosum CH08 grown in PDB for 48 h. And when 0.7 M sucrose was used as osmotic stabilizer in the regeneration medium OPDA, the regeneration efficiency of protoplasts was 41.48%. Thirdly, the mutagenesis conditions and secondary screening model of C. cupreum CH21 were explored. An 80% to 85% death rate could be achieved when the protoplasts of C. cupreum CH21 were irradiated by 15 w UV lamp from 30 cm distance for 90 s. In addition, the doublelayer plate’s method for the primary screening of high-producing antibiotics strains was established. A high yielding antibiotic mutant CH21-I-402 was obtained through the primary screening on plate and the secondary selection in Erlenmeyer flask, compared to the original CH21 strain, the antifungal activity of the mutant CH21-I-402 was increased by 18.3%. Fourth, the sensitivity to antibiotics of both C. cupreum CH21-I-402 and C. globusm CH08 was detected. Results showed C. cupreum CH21-I-402 was sensitive to G418 (Geneticin) (Gs) and resistant to Hygromycin B(Hr), and C. globusm CH08 was sensitive to both G418 (Geneticin) (Gs) and Hygromycin B(Hs). At the same time, neomycin phosphotransferase II (npt II) gene was transformed into C. globusm CH08(Gs, Hs) mediated by Agrobacterium tumefaciens EHA105, and the npt II gene was verified by polymerase chain reaction in resistance to G418 strains CH08-GR70 and CH08-GR120. The transformants still showed sensitive to Hygromycin B(Hs). Finally, a selection system for hybrids was set up by interspecies protoplast fusion between C. cupreum and C. globusm using dominant selective drug resistance markers. At first, protoplasts of C. cupreum CH21-I-402 (Hr, Gs) and C. globusm CH08-GR70 (Hs, Gr) were prepared, then the protoplasts were fused in the presence of 35% polyethylene glycol 6000 and regenerated on OPDA medium with 65 μg/ml Hygromycin B and 60μg/ml G418, at last 46 colonies with Hr and Gr were obtained. Even after 5 generations’ subculture, most of the colonies displayed significant difference in taxonomic characteristics with their parental strains. Regenerated strains PF1 and PF26 were confirmed as fusants by amplified fragment length polymorphisms analysis with the genomic DNA as the model. PF26 showed higher inhibitory activity against Colletotrichum gloeosporioides and Macrophoma kuwatsukai than that of the parental strain C. globusm.

一氧化氮对杨树耐旱性的影响

探讨了外源NO对水分胁迫下杨树叶片质膜相对透性、叶片光合作用和氧化伤害保护酶的影响.结果表明,NO供体硝普钠(sodiumnitroprusside,SNP)能提高杨树叶片的含水率,在水分胁迫(PEG6000渗透液处理)下,能缓解叶片的水分丢失.NO对杨树叶片光合作用具有双重性,低浓度SNP(200、500μmol·L-1)能促进叶片的光合,高浓度SNP(1000、2000μmol·L-1)则明显抑制叶片的光合.较短时间水分处理胁迫(1h))的杨树叶片SOD和POD活性显著高于较长时间(3h)水分胁迫下叶片的酶活性.经SNP处理后,各处理组POD、SOD活性明显上升.同时,随SNP浓度的增加,POD和SOD活性表现出先上升后下降的趋势.外源NO可通过诱导POD和SOD活性的上升,延缓活性氧的积累,从而减轻水分胁迫对杨树的伤害,增强树木的耐旱能力.

Studies on water-swellable elastomers .2. Thermal properties and crystalline behavior of amphiphilic graft copolymers with poly(ethylene glycol) side chains

The thermal properties and crystalline structure of the amphiphilic graft copolymers CR-g-PEG600, CR-g-PEG2000, and CR-g-PEG6000 using chloroprene rubber (CR) as the hydrophobic backbone and poly(ethylene glycol) (PEG) with different molecular weights as the hydrophilic side chains were studied by DSC and WAXD. The results showed that a distinct phase-separated structure existed in CR-g-PEGs because of the incompatibility between the backbone segments and the side-chain segments. For all the polymers studied, T-m2, which is the melting point of PEG crystalline domains in CR-g-PEG, decreased compared to that of the corresponding pure PEG and varied little with PEG content. For CR-g-PEG600 and CR-g-PEG2000, T-m1, which is the melting point of the CR crystalline domains, increased with increasing PEG content when the PEG content was not high enough, and at constant PEG content, the longer were the PEG side chains the higher was the T-m1. The crystallite size L-011 of CR in CR-g-PEGs increased compared to that of the pure CR and decreased with increasing PEG content. (C) 1997 John Wiley & Sons, Inc.

抗旱性不同的春小麦品种籽粒萌发期α-淀粉酶活性及其同工酶分析

为了解春小麦萌发期生理生化变化对干旱胁迫的响应及为早期抗旱筛选鉴定提供科学依据,选用抗旱性不同的9个春小麦品种为材料,在20%聚乙二醇(PEG6000)干旱胁迫下和非干旱胁迫下进行萌发试验,研究了α-淀粉酶活性及其同工酶的表达。结果表明:1)在两种胁迫处理下,α-淀粉酶活性在品种间都存在着差异,但干旱胁迫下抗旱品种α-淀粉酶活性显著高于干旱敏感品种;2)α-淀粉酶活性与胚芽鞘长度之间呈显著正相关;3)在20%PEG6000胁迫下,抗旱品种α-淀粉酶同工酶受抑制较小,条带较多,胚芽鞘长度与主胚根长度受抑制较小。因此认为,抗旱品种在干旱胁迫下有着较高的萌发势,可能与具有表达α-淀粉酶同工酶的强势基因型有关;干旱胁迫下α-淀粉酶活性和α-淀粉酶同工酶可以作为春小麦抗旱性筛选和鉴定的指标。

Evaluating the effects of tannins on the extent and rate of in vitro measured gas and methane production using the Automated Pressure Evaluation System (APES)

An in vitro study was conducted to investigate the effect of tannins on the extent and rate of gas and methane production, using an automated pressure evaluation system (APES). In this study three condensed tannins (CT; quebracho, grape seed and green tea tannins) and four hydrolysable tannins (HT; tara, valonea, myrabolan and chestnut tannins) were evaluated, with lucerne as a control substrate. CT and HT were characterised by matrix assisted laser desorption ionisation-time of flight mass spectrometry (MALDI-TOF-MS). Tannins were added to the substrate at an effective concentration of 100 g/kg either with or without polyethylene glycol (PEG6000), and incubated for 72 h in pooled, buffered rumen liquid from four lactating dairy cows. After inoculation, fermentation bottles were immediately connected to the APES to measure total cumulative gas production (GP). During the incubation, 11 gas samples were collected from each bottle at 0, 1, 4, 7, 11, 15, 23, 30, 46, 52 and 72 h of incubation and analysed for methane. A modified Michaelis-Menten model was fitted to the methane concentration patterns and model estimates were used to calculate the total cumulative methane production (GPCH4). GP and GPCH4 curves were fitted using a modified monophasic Michaelis-Menten model. Addition of quebracho reduced GP (P=0.002), whilst the other tannins did not affect GP. Addition of PEG increased GP for quebracho (P=0.003), valonea (P=0.058) and grape seed tannins (P=0.071), suggesting that these tannins either inhibited or tended to inhibit fermentation. Addition of quebracho and grape seed tannins also reduced (P≤0.012) the maximum rate of gas production, indicating that microbial activity was affected. Quebracho, valonea, myrabolan and grape seed decreased (P≤0.003) GPCH4 and the maximum rate (0.001≤ P≤ 0.102) of CH4 production. Addition of chestnut, green tea and tara tannins did not affect total gas nor methane production. Valonea and myrabolan tannins have most promise for reducing methane production as they had only a minor impact on gas production.

Effect of water-soluble polymers, polyethylene glycol and poly(vinylpyrrolidone),on the gelation of aqueous micellar solutions of Pluronic copolymer F127

The micellization of F127 (E98P67E98) in dilute aqueous solutions of polyethylene glycol (PEG6000 and PEG35000) and poly(vinylpyrrolidone) (PVP K30 and PVP K90) is studied. The average hydrodynamic radius (rh,app) obtained from the dynamic light scattering technique increased with increase in PEG concentration but decreased on addition of PVP, results which are consistent with interaction of the micelles with PEG and the formation of micelles clusters, but no such interaction occurs with PVP. Tube inversion was used to determine the onset of gelation. The critical concentration of F127 for gelation increased on addition of PEG and of PVP K30 but decreased on addition of PVP K90. Small-angle X-ray scattering (SAXS) was used to show that the 30 wt% F127 gel structure (fcc) was independent of polymer type and concentration, as was the d-spacing and so the micelle hard-sphere radius. The maximum elastic modulus (G0 max) of 30 wt% F127 decreased from its value for water alone as PEG was added, but was little changed by adding PVP. These results are consistent with the packed-micelles in the 30 wt% F127 gel being effectively isolated from the polymer solution on the microscale while, especially for the PEG, being mixed on the macroscale.

Hydric restriction as vigor assessment method of soybean seeds

The objective of this study was to investigate the possibility of using hydric restriction as a method for evaluating vigor of soybean seeds. The soybean seeds, cultivar BRS 245RR, represented by four different seed lots, were characterized by germination and vigor. For the treatment of hydric restriction and temperature, the combination of substrate water potential and temperature were the following: deionized water (0.0 MPa); polyethylene glycol (PEG 6000) aqueous solution (-0.1, -0.3 and -0.5 MPa); and four temperatures (20 ºC, 25 ºC, 30 ºC, and 35 ºC), respectively. A completely randomized experimental design was used, with four replications per treatment, and the ANOVA was performed individually for each combination of temperature and water potential of substrate. According to results obtained, the test of hydric restriction has the same efficiency of the accelerated aging test in estimating vigor of soybean seeds, cv. BRS 245RR, when water potentials of -0.1 MPa or -0.3 MPa at a temperature of 25 ºC, or -0.3 MPa at a temperature of 30 ºC are used.

Análise in vitro da expressão de genes de resposta a estresse oxidativo e osmótico da bactéria fastidiosa Leifsonia xyli subsp. xyli

Atualmente, o Brasil é o maior produtor de cana-de-açúcar (Saccharum ssp.), no qual o estado de São Paulo é responsável por mais de 50% da produção. Esta cultura é hospedeira de diversos patógenos que podem limitar sua produção, dentre os quais se destaca a bactéria Leifsonia xyli subsp. xyli (Lxx), agente causal do raquitismo da soqueira (ratoon stunting disease - RSD). Pouco se sabe sobre a fisiologia deste organismo e quais as estratégias utilizadas por este para colonizar seu hospedeiro. No entanto, sabemos que para infectar e colonizar seus hospedeiros, é necessário que bactérias parasíticas superem estresses de diversas naturezas impostas durante estes processos, como os estresses oxidativo e o osmótico. Neste contexto, os objetivos deste trabalho foram identificar in silico e analisar a expressão in vitro, por qPCR, de genes relacionados a estes dois estresses. Uma análise da sequência do genoma de Lxx identificou 35 genes, sendo 8 relacionados ao estresse oxidativo, 9 relacionados ao estresse osmótico e 11 relacionados a estresse gerais, incluindo um cluster de 6 genes envolvidos na síntese de carotenoides. A expressão destes foi avaliada 60 minutos após exposição a 30mM de H2O2 ou 7% (p/v) de polietilenoglicol 6000 (PEG 6000). Sete genes foram avaliados como normalizadores das reações de qPCR. A quantificação do grau de peroxidação lipídica indicou que ambos os tratamentos resultaram em sensível peroxidação, muito embora o efeito do tratamento com PEG 6000 tenha sido maior do que o tratamento com H2O2. A exposição ao H2O2 aumentou a expressão dos genes katA (catalase), sodA (superóxido dismutase), msrA (Sulfóxido de metionina redutase) e msrB (Sulfóxido de metionina redutase) bem como de todos os genes responsáveis pela síntese de carotenoides. Por outro lado, todos os genes relacionados ao estresse osmótico foram menos expressos na presença deste composto. Já quando a bactéria foi exposta a PEG 6000, o oposto ocorreu, ou seja, os genes relacionados ao estresse osmótico, que são otsA (Trealose-6-fosfato sintase), otsB (Trealose fosfatase), treY (Malto-oligosil trealose sintase), treZ (Malto-oligosil trealose trealoidrolase), treS (Trealose sintase), proX (Proteína de ligamento em substrato, tipo ABC glicina betaína transportadora), proW (Proteína permease, tipo ABC glicina betaína transportadora), proZ (Proteína permease, tipo ABC glicina betaína transportadora) e Naggn (Amidotransferase), além dos genes do cluster carotenoide, foram mais expressos, ao passo que alguns dos genes ligados à resposta ao estresse oxidativo foram menos expressos. Verificou-se também, através de PCR convencional utilizando primers para amplificar as regiões entre os genes carotenoides, que estes são expressos como um RNA policistrônico, constituindo assim um operon. Estes resultados validam predições anteriores baseadas na análise in silico da sequência do genoma de Lxx, confirmando que Lxx possui mecanismos responsivos aos estresses osmótico e oxidativo aos quais é submetida durante o processo de infecção de seu hospedeiro.