[Objective] The purpose of this study was to clone a starch phosphorylase gene from Dunaliella salina and to preliminarily analyze its basic properties and protein expression. [Method] RT-PCR and RACE （rapid amplific...[Objective] The purpose of this study was to clone a starch phosphorylase gene from Dunaliella salina and to preliminarily analyze its basic properties and protein expression. [Method] RT-PCR and RACE （rapid amplification of cDNA ends） method was used for gene cloning; basic properties of the gene were analyzed using bioinformatics method; prokaryotic expression vector PGS21a-DsSP was constructed and transformed into E. coil BL21; the fusion protein was purified and detected by GST-SefinoseTM Kit and Western Blot, respectively. [Result] A starch phos-phorylase gene （GenBank accession No. KF061044） named DsSP was successfully isolated from D. salina. Basic properties, subcellular localization, secondary structure and tertiary structure of the protein were analyzed and predicted. The fusion protein was found in the supernatant and inclusion bodies. The supernatant protein was successfully purified. Western Blot analysis showed that the fusion protein was successfully expressed in E. coil BL21. [Conclusion] This study laid experimental foun- dation for further clarifying the function and mechanism of DsSP.展开更多
三阴性乳腺癌(Triple-Negative Breast Cancer,TNBC)是恶性程度较高的肿瘤之一,由于其常规化疗效果不佳,且缺乏有效的靶向治疗方案,TNBC仍然是一种预后差、治疗方案选择少的疾病。抗肿瘤血管生成疗法作为癌症治疗的重要方法之一,目前抗...三阴性乳腺癌(Triple-Negative Breast Cancer,TNBC)是恶性程度较高的肿瘤之一,由于其常规化疗效果不佳,且缺乏有效的靶向治疗方案,TNBC仍然是一种预后差、治疗方案选择少的疾病。抗肿瘤血管生成疗法作为癌症治疗的重要方法之一,目前抗肿瘤血管生成药物在临床肿瘤治疗中已广泛应用。研究表明,TNBC中存在包括血管生成因子(Vascular endothelial growth factor,VEGF)在内的血管新生关键基因的异常表达,抗血管生成疗法在TNBC的临床治疗中具有较大潜力,且研究发现抗血管生成疗法对于TNBC的治疗结果有显著的进展,可以作为治疗TNBC的有效治疗手段,现对抗血管生成药物在TNBC治疗中的应用及研究进展作一综述。展开更多
The key limiting factors to high-density culture of Porphyridium cruentum are the uptake of light energy and nutrient by the microalgal cells. Under the optimal conditions of carrier culture, both cell mass and cell d...The key limiting factors to high-density culture of Porphyridium cruentum are the uptake of light energy and nutrient by the microalgal cells. Under the optimal conditions of carrier culture, both cell mass and cell density were increased significantly up to 5.2 g/L (DW) and 5.2107/ml. Furthermore, the effects of the liquid circulation velocity, light intensity and initial cell density on cell mass productivity of P. cruentum were investigated in a 42 L internal loop airlift photobioreactor. Although the light intensity was as low as 100 mmol/(m2s), the light damage or the photoinhibition phenomenon was observed under the culture condition of low initial cell mass (0.10 g/L, DW) and high liquid circulation velocity (0.30 m/s). However, a higher cell growth rate and a high cell mass productivity were achieved with the same conditions only at high initial cell mass (about 0.80 g/L, DW). Under the optimal conditions, the cell specific growth rate, cell mass volumetric and areal output rate reached to 0.95 d-1, 0.80 g/(Ld) and 42.5 g/(m2d) respectively. Finally, a method of nutrient feeding and gradual increase of light intensity in different cultural stages was developed, which further improved the cell mass, cell mass volumetric and areal output rate to 5.9 g/L, 1.2 g/(Ld) and 61.7 g/(m2d) respectively.展开更多
基金Supported by National Natural Science Foundation of China(No.30972240)Science and Technology Project of Liaoning Provincial Department of Education(No.2008T023)~~
文摘[Objective] The purpose of this study was to clone a starch phosphorylase gene from Dunaliella salina and to preliminarily analyze its basic properties and protein expression. [Method] RT-PCR and RACE （rapid amplification of cDNA ends） method was used for gene cloning; basic properties of the gene were analyzed using bioinformatics method; prokaryotic expression vector PGS21a-DsSP was constructed and transformed into E. coil BL21; the fusion protein was purified and detected by GST-SefinoseTM Kit and Western Blot, respectively. [Result] A starch phos-phorylase gene （GenBank accession No. KF061044） named DsSP was successfully isolated from D. salina. Basic properties, subcellular localization, secondary structure and tertiary structure of the protein were analyzed and predicted. The fusion protein was found in the supernatant and inclusion bodies. The supernatant protein was successfully purified. Western Blot analysis showed that the fusion protein was successfully expressed in E. coil BL21. [Conclusion] This study laid experimental foun- dation for further clarifying the function and mechanism of DsSP.
基金Supported by the China Ministry of Science and Technology(No. 96-C02-04-05)
文摘The key limiting factors to high-density culture of Porphyridium cruentum are the uptake of light energy and nutrient by the microalgal cells. Under the optimal conditions of carrier culture, both cell mass and cell density were increased significantly up to 5.2 g/L (DW) and 5.2107/ml. Furthermore, the effects of the liquid circulation velocity, light intensity and initial cell density on cell mass productivity of P. cruentum were investigated in a 42 L internal loop airlift photobioreactor. Although the light intensity was as low as 100 mmol/(m2s), the light damage or the photoinhibition phenomenon was observed under the culture condition of low initial cell mass (0.10 g/L, DW) and high liquid circulation velocity (0.30 m/s). However, a higher cell growth rate and a high cell mass productivity were achieved with the same conditions only at high initial cell mass (about 0.80 g/L, DW). Under the optimal conditions, the cell specific growth rate, cell mass volumetric and areal output rate reached to 0.95 d-1, 0.80 g/(Ld) and 42.5 g/(m2d) respectively. Finally, a method of nutrient feeding and gradual increase of light intensity in different cultural stages was developed, which further improved the cell mass, cell mass volumetric and areal output rate to 5.9 g/L, 1.2 g/(Ld) and 61.7 g/(m2d) respectively.
