蓝图转白图的关键技术研究

在工程勘察设计方面,工程建设项目使用及业主单位归档的图纸均是由设计单位提供的蓝图,设计单位归档的图纸是底图和蓝图。然而蓝图时间久了就会变黄、褪色、沾水即模糊,不易长期保存,再加上会排放出氨气或显影液的气味对环境污染极其严重,影响从业人员的职业健康,导致工作效率低下,人员密集,成本过高的弊端日益显露。因此,寻找蓝图的替代物已成为人们关注的课题。本文对"蓝"转"白"的演变背景、本质以及关键技术进行了探讨,仅供行业参考。

水稻白条纹转绿突变体wsl887的鉴定和基因定位

温敏型水稻叶色突变基因的挖掘可以丰富水稻遗传资源,且为研究低温调控叶绿体发育机制的材料。本研究利用;Co射线辐射诱变贵州地方稻种资源桥港珍珠米,获得一个新的白条纹转绿突变体wsl887。通过表型鉴定和主要农艺性状调查可知,在自然条件下wsl887叶幼苗呈现白条纹表型,成熟后主要农艺性状与野生型相比均无显著差异;在20℃条件下wsl887叶片呈现白化,光合色素含量显著降低;在25℃条件下wsl887叶片呈现白色条纹,光合色素含量显著升高;在30℃条件下wsl887叶色与野生型无明显差异,光合色素含量无显著差异。透射电镜观察显示,在20℃条件下,wsl887叶肉细胞中无叶绿体或者叶绿体发育畸形;在30℃条件下,wsl887叶绿体数量和形态均恢复正常。qRT-PCR分析表明,wsl887在20℃和30℃条件下,与野生型相比,部分光合色素代谢途径基因、光合作用及线粒体电子传递相关基因的表达水平呈现不同程度的变化。遗传分析表明,wsl887突变体的表型受一对隐性核基因WSL887控制,该基因被定位于2号染色体上的SSR标记RM262和RM5427之间,物理距离为743.6 kb。该白条纹转绿突变体wsl887是一个新的温敏型叶色突变体,在自然条件下其主要农艺性状没有发生显著变化,未对植株产量造成影响,因此在杂交水稻上具有较大的应用价值。在不同温度条件下,由细胞质编码的基因表达量均显著减少,由此推测该突变基因可能与细胞质中叶绿体的发育有关。本研究结果为进一步克隆wsl887突变基因和研究基因功能奠定了一定的理论基础。

基于气相色谱-质谱联用技术的转人血清白蛋白水稻的代谢组学方法研究

为推动代谢组学研究,利用转人血清白蛋白水稻,对样品前处理、色谱柱选择、GC-MS的检测方法、谱图处理和数据分析等方面进行研究。结果显示:在提取液为水∶乙腈∶异丙醇=1∶1.5∶1.5的条件下,30℃衍生90min,作为样品前处理条件,选用DB-5MS色谱柱,采用保留时间锁定的数据采集方法,用AMDIS软件的保留时间分析模式对谱图解卷积,然后经过MPP软件进行主成分分析,建立了一套完整的代谢组学研究方法流程。

白酒产业转型升级路径分析研究——以泸州为例

白酒产业是泸州市特色优势产业,为泸州经济社会发展做出了重要贡献。进入“十四五”时期,泸州市白酒产业面临白酒产业政策调整、新冠肺炎疫情、中美贸易争端以及中国加快构建“双循环”新发展格局等机遇和挑战。为探究新形势下泸州市白酒制造业转型升级路径,本研究通过梳理文献、深入调研、座谈等方法,从行业发展的角度入手,系统梳理相关文献,对泸州白酒产业现状、面临形势进行了分析,并在此基础上提出了创新发展、绿色发展、融合发展三个方面的建议。

ZNT1 Involves Cuproptosis through Regulating MTF1-conduced Expression of MT1X under Copper Overload

Industrial activities such as smelting emissions,mineral combustion and industrial wastewater discharge might lead to copper pollution in the environment.This kind of copper pollution has harmful effects on aquatic o rganisms,plants and animals through direct or indirect exposure.However,the current understanding of the toxicity of copper is rather limited.Copper overload can perturb intracellular homeostasis and induce oxidative stress and e ven cell death.Recently,cuproptosis has been identified as a copper-dependent form of cell death induced by o xidative stress in mitochondria.We uncover here that zinc transporter 1(ZNT1)is an important regulator involved in cuproptosis.Firstly,we established the copper overload-induced cell death model with the overexpression of copper importer SLC31A1 in HeLa cells.Using this model,we conducted unbiased genome-wide CRISPR-Cas9 screens in cells treated with copper.Our results revealed a significant enrichment of ZNT1 gene in both library A and library B plasmids.Knocking out of ZNT1 in HeLa cells notably prevented cuproptosis.Subsequent knockout of metal transcription factor 1(MTF1)in ZNT1-deficient cells nearly abolished their ability to resist copper-induced cell death.However,overexpression of metallothionein 1X(MT1X)in the double-knockout cells could p artially restored the resistance to cuproptosis by loss of MTF1.Mechanistically,knockout of ZNT1 could promote MT1X expression by activating MTF1.As a consequence,the interaction between MT1X and copper was e nhanced,reducing the flow of copper into mitochondria and eliminating mitochondria damage.Taken together,this study reveals the important role of ZNT1 in cuproptosis and shows MTF1-MT1X axis mediated resistance to c uproptosis.Moreover,our study will help to understand the regulatory mechanism of cellular and systemic copper homeostasis under copper overload,and present insights into novel treatments for damages caused by both genetic copper overload diseases and environmental copper contamination.

骨髓增生异常综合征转化为急性白血病的高危因素分析

目的:探讨影响骨髓增生异常综合征(MDS)转化为急性白血病的高危因素。方法:回顾性分析我院收治的101例MDS患者的转白时间、转白亚型、临床特征、实验室数据等临床资料。结果:101例MDS患者中,34.6%(35/101)转化为急性髓系白血病,中位转白时间7.5(0.5~53)个月,中位生存时间40.7(1~87)个月。单因素分析发现,染色体核型、骨髓原始细胞比例、血细胞减少系数、病态造血累及系数、WHO分型均影响MDS患者的转白率(均P<0.05)。多因素分析发现,染色体核型、病态造血系数、WHO分型是MDS向急性白血病转化的独立危险因素(均P<0.05)。转白组患者较未转白组患者的生存期明显缩短(11个月∶45个月)。结论:MDS是一类高风险向急性白血病转化的疾病,根据相关高危因素,对其进行预后评估,从而为个体化治疗提供临床依据,对延长患者转白时间及总生存时间具有重要意义。

Engineering The Neck Hinge Reshapes The Processive Movement of Kinesin-3

Objective In kinesin-3,the neck coil correlates with the following segments to form an extended neck that contains a characteristic hinge diverse from a proline in KIF13B to a long flexible linker in KIF1A.The function of this neck hinge for controlling processive movement,however,remains unclear.Methods We made a series of modifications to the neck hinges of KIF13B and KIF1A and tested their movement using a single-molecule motility assay.Results In KIF13B,the insertion of flexible residues before or after the proline differentially impacts the processivity or velocity,while the removal of this proline increases the both.In KIF1A,the deletion of entire flexible neck hinge merely enhances the processivity.The engineering of these hinge-truncated necks of kinesin-3 into kinesin-1 similarly boosts the processive movement of kinesin-1.Conclusion The neck hinge in kinesin-3 controls its processive movement and proper modifications tune the motor motility,which provides a novel strategy to reshape the processive movement of kinesin motors.

Pathogen_resistant Transgenic Plant of Brassica pekinensis by Transfering Antibacterial Peptide Gene and Its Genetic Stability

The soft rot infected by pathogenic bacterium Erwinia aroideae Holland is one of the three serious diseases of Chinese cabbage ( Brassica pekinensis Rupr.). By constructing vector system of high frequency transformation mediated by Agrobacterium tunefaciens EHA105, anti-bacterial peptide gene with strong bactericidal action to pathogenic bacteria was introduced into Chinese cabbage AB-81 self-bred line and the transgenic plants were obtained. PCR and Southern blotting detection showed that target gene was integrated into plant genome of Chinese cabbage. The tests of bacteriostasis action of the extract from transgenic plants in vitro, and the assay of disease-resistant of transgenic plantlets in the tube and the pot by perfusing inoculation with pathogenic bacteria showed obvious resistance to soft rot. This resistance can be a stable heredity by genetic analysis of generations of transgenic plants self-bred, separation ratio of its R, was 3:1. The resistance to Km and disease of soft rot was still kept in the R-5. These results indicated the possibility of breeding new varieties of anti-soft rot Chinese cabbage by transgenic plants as parents.

Cloning and Characterization of the Na^+/H^+ Antiport Genes from Triticum aestivum

The Na+/H+ antiport genes namedTaNHX1andTaNHX2were cloned by screening a salt_stressed wheat cDNA library using rice Na+/H+ antiport cDNA fragment as the probe. Sequencing analysis showed thatTaNHX1was 2 029 bp in length and contained a complete ORF of 1 638 bp. TheTaNHX1encodes a polypeptide of 546 amino acids with a transmembrane domain DIFFIYLLPPI.TaNHX2was 1 693 bp in length consisting of a partial ORF followed by a 3′_UTR of 808 bp. The amino acid sequence of these two genes were about 70% identical to the known NHX genes from rice, Arabidopsis and Atriplex. A RT_PCR assay showed that the level ofTaNHX1transcripts was increased and reached a steady higher level in the seedlings after 3 h treatment with 400 mmol/L NaCl.

Physiological Character and Gene Mapping in a New Green-revertible Albino Mutant in Rice

A green-revertible albino mutant-Qiufeng M was found from the japonica rice （Oryza sativa L. ssp. japonica） Qiufeng in the field. The first three leaves of the mutant were albino with some green. The leaf color became pale green since the fourth leaf and the glume had the same phenomenon as the first three leaves. The measuring data of the pigment content confirmed the visually observed results. It truly had a remarkable changing process in the leaf color in Qiufeng M. Comparison of the main agronomic characters between Qiufeng and Qiufeng M indicated that the neck length and grain weight showed significant difference at the 1% level, and other characters were not different. Genetic analysis showed that the green-revertible albino trait was controlled by a single recessive nucleic gene. Using 209 recessive mutant individuals in the F2 population derived from the cross Pei＇ai 64S × Qiufeng M, a gene, tentatively named gra（t）, was located between the SSR markers of RM475 and RM2-22 on the long arm of chromosome 2. The genetic distance were 17.3 cM and 2.9 cM respectively.

Human Pro-insulin Transgenic Calf Derived from Somatic Cell Nuclear Transfer

The current study was undertaken to evaluate the possibility of producing a human pro-insulin transgenic cow by means of somatic cell nuclear transfer （SCNT）. A double selection system, Neomycin resistance （Neo^r） gene and enhanced green fluorescent protein （EGFP） gene linked through an inner ribosomal entry site （IRES） sequence directed by a Cytomegalovirus （CMV） promoter, was used for enrichment and selection of the transgenic cells and preimplantation embryos. Transgenes were introduced into bovine fetal fibroblast cells （BFF） cultured in vitro through electroporation （900 V/cm, 5 ms）. Transgenic bovine fibroblast cells （TBF） were enriched through addition of G418 in culture medium （800 μg/mL）. Before being used as a nuclear donor, the TBF cells were either cultured in normal conditions （10% FBS） or treated with serum starvation （0.5% FBS for 2-4 days） followed by 10 hours recovery for G1 phase synchronization. Transgenic cloned embryos were produced through GFP-expressing cell selection and SCNT. The results were the percentage of blastocyst development following SCNT was lower using TBF than BFF cells （23.2% VS 35.2%, P 〈 0.05）. No difference in the percentage of cloned blastocysts between the two groups of transgenic nuclear donor of normal and starvation cultures were observed （23.2% VS 18.9%, P 〉 0.05）. Two to four GFP-expressing blastocysts were transferred into the uterus of each synchronised recipient. One pregnancy from of seven recipients （21 embryos） was confirmed by rectum palpation 60 days after embryo transfer and one recipient has given birth to a calf at term. PCR and DNA sequencing analysis confirmed that the calf was produced using human proinsulin transgenic animal.

Novel Halophyte EREBP/AP2-type DNA Binding Protein Improves Salt Tolerance in Transgenic Tobacco

EREBP/AP2-type proteins are members of a large DNA binding protein (DBP) family found in plants. Some members like APETALA2 and AtDREB/CBF can regulate flower development and response to environmental stresses, respectively. To characterize transcription factors involved in plant responses to salt stress, we constructed cDNA library from salt-treated halophyte (Atriplex hortensis) and isolated a novel gene encoding EREBP/AP2-type protein from this library. This cDNA contained an ORF of 723 bp and a long 3'-Untranslated-Region (UTR) of 655 bp. The deduced amino acid sequence showed one conserved DNA binding domain of EREBP/AP2, thus the corresponding gene was named AhDREB1 with a calculated molecular mass of 26.1 kD. AhDREB1 under the control of CaMV 35S promoter was then transformed into tobacco and nine independent transgenic lines were obtained and subjected to long term salt stress. The results suggested that overexpression of AhDREB1 improved the salt tolerance in transgenic tobacco through functioning as a regulatory molecule in response to salt stress. Analysis of Arabidopsis genome in database resulted in dozens of EREBP/AP2-type homologous proteins, of which seven members showed high similarity to AhDREB1. Secondary structure analysis predicted similar arrangement of a-helix in their DNA binding domains.

Improvement of the Hydroponic Growth and Waterlogging Tolerance of Petunias by the Introduction of vhb Gene

The coding sequence of Vitreoscilla hemoglobin (vhb) was cloned with PCR technique from Vitreoscilla stercoraria Pringsheim. The plant expression vector with vhb gene under the control of CaMV 35S promoter was constructed and used in the transformation of Petunia hybrida Vilm by the Agrobacterium mediated procedure. The results of PCR amplification and Southern hybridization indicated that the vhb gene had been integrated into the petunia genome and the vhb gene expression had been detected by RT-PCR amplification. In hydroponic culture the transgenic petunias grew much better than non-transgenic controls. For further analysis of hypoxia tolerance of transgenic petunia, the petunia plants with vhb gene were submerged into liquid MS medium. The transgenic plants survived in hypoxic condition and grew out of the liquid surface in a few weeks, while non-transgenic plants were still submerged and suffocated in culture solution without ability to grow out of liquid medium in submersed culture for four to five weeks. The vhb gene transformed petunia plants had been planted and tested in a simulated flooding condition, and showed obvious tolerance to water-logging. It seen is that hemoglobin gene from Vitreoscilla might have the potential use in molecular breeding for the improvement of plant resistance to hypoxia and flooding.

Screening an Na^+/H^+ Antiporter Gene from the Halophiles Colonizing in the Dagong Ancient Brine Well of Zigong City,China

[Objective] This study aimed to screen an Na＋/H＋ antiporter gene from the halophiles colonizing in the Dagong Ancient Brine Well in Zigong City, China, and then analyze the gene structure and properties of the protein encoded by this gene. [Method] Metagenomic DNA libraries of halophiles from the Dagong Ancient Brine Well were used for screening genes with Na＋/H＋ antiporter activity in antiporter-defi- cient E. coil KNabc strain by functional complementation. Then the start codon, stop codon, ORF, -35 region, -10 region and SD sequence of Na~/H＋ antiporter gene, as well as the molecular weight, isoelectric point, hydrophobic region, transmembrane domain, phyletic evolution and salt resistance of protein encoded by the gene were investigated. [Result] A new Na＋/H＋ antiporter gene m-nha was obtained, which ,ren- dered the antiporter-negative mutant E. coil KNabc cells with both the resistance to Na＋ and the ability to grow under alkaline conditions. [Conclusion] The structure and amino acid sequence of M-Nha was different from the previously reported Na＋/H~ antiporters, and the m-nha gene disclosed from the Dagong Ancient Brine Well was identified as a novel Na＋/H＋ antiporter gene. This study was significant not only in helping us understand the salt tolerance of halophiles in ancient brine wells and develop and utilize the genes resource, but also in exploring new salt-tolerant genes.

Overexpression of Proline Transporter Gene Isolated from Halophyte Confers Salt Tolerance in Arabidopsis

Proline is one of the most important and widespread osmolyte which functions in adaptation to adverse environmental stresses in many organisms. Also it is an important carbon and nitrogen resource in higher plants. Metabolism of proline has been elucidated in many plant species. However, transport of proline was poorly characterized although transport system plays an important role in proline distribution in different tissues. We isolated one full_length cDNA encoding proline transporter from the typical halophyte: Atriplex hortensis L. through cDNA library screening and 5′_RACE. The deduced amino acid sequence had eleven transmembrane domains, showed 60%-69% similarities to other ProTs and the gene was designated AhProT1. In the phylogenetic tree, higher plants' ProTs, e.g. AhProT1, showed more similar to ProP from microorganisms than ProT from mammalians. AhProT1 gene was transformed into Arabidopsis thaliana under 35S promoter. In MS medium containing [U_ 14 C] proline, AhProT1 + plants were able to accumulate much more radiolabeled proline in the roots than control plants. In MS medium containing different concentrations of NaCl, AhProT1 + plants could endure 200 mmol/L NaCl and keep development and biomass increase with proline supply, whereas control plants died back at 150 mmol/L NaCl.

Studies of Transgenic Hybrid Poplar 741 Carrying Two Insect-resistant Genes

Partially modified Bt Cry1Ac gene and the arrowhead proteinase inhibitor (API) gene were used to construct a plant transformation vector pBtiA and this construct was transferred into the genome of the hybrid poplar 741 [ Populus alba L.×( P. davidiana Dode+ P. simonii Carr.)× P. tomentosa Carr.] by Agrobacterium _ mediated transformation. Ten kanamycin resistant plants have been regenerated. Upon insect bioassay using Clostera anachoreta (Fabricius), three of the examined plants were demonstrated to be highly resistant to the testing insects. The mortality of insect larvae on one plant was higher than 90% in 6 days after infestation and the growth of the survival larvae were seriously inhibited. Results of PCR and Southern blot analysis indicated that both Bt Cry1Ac gene and API gene were integrated as a single copy into the genomes of these three plants when Cry1Ac gene fragment was used as the probe. Protein dot blot immunoassay and ELISA analysis revealed that at least the Cry1Ac protein was produced in these three transgenic plants and the expression levels were estimated to be approximately 0.015% of the leaf total soluble protein. This is the first report on insect resistant transgenic hybrid poplar 741 that expresses two insecticidal protein genes.

Virus Movement Protein Gene Mediated Resistance Against Cucumber Mosaic Virus Infection

Tobacco ( Nicotiana tabacum L.) “NC89” plants were transformed with deletion mutant of cucumber mosaic virus (CMV) movement protein (MP) gene and full_length CMV MP gene, respectively. The transformed plants were analyzed with polymerase chain reaction (PCR), PCR_Southern, Southern and Western blots. R 0 generation of the transgenic plants were inoculated with CMV. Five out of 10 lines of tobacco plants (BMPK) transformed with CMV MP deletion mutant gene showed high resistance to CMV infection and remained symptomless for up to 50 days post_inoculation. In contrast, tobacco plants (BMPR) transformed with full_length CMV MP gene did not show resistance to CMV infection. However, most of the infected full_length CMV MP gene transgenic plants recovered by showing none or very mild mosaic symptoms in 40 days post_inoculation. The results of R 1 generation of the BMPK transgenic plants tested under field conditions showed that all 5 lines of transgenic plants could delay the virus disease development.

Effects of two kinds of transgenic poplar on protective enzymes system in the midgut of larvae of American white moth

The leaves of Bt (Bacillus thuringiensis) transgenic poplar (Populus nigra L.) and CpTI (Cowpea trypsin inhibitor) transgenic poplar ((P. tomentosa×P. bolleana)×P. Tomentosa) were taken to feed the 4th-5th-instar larvae of American white moth (Hyphantria cunea (Drury)) for determination of the activities of the protective enzyme system inside larvae’s body. The physiological and biochemical effects of the transgenic poplars on the larvae were studied. The results showed that the two kinds of transgenic poplars had similar effects on the protective enzyme system in the midgut of larvae. The activities of superoxide dismutase, catalase, and peroxidase in midgut of the larvae increased gradually, reached the highest value at a certain time, and then decreased suddenly. For the larvae that were fed with the leaves of Bt transgenic poplar, the peak value of superoxide dismutase and catalase presented at the time of 24-h feeding, while the peak of peroxidase took place at the time of 12-h feeding. The activities of these protective enzymes for the larvae that were fed with leaves of CpTI transgenic poplar peaked 12 h later than that of those fed with leaves of Bt transgenic poplar. The comparison of activities of the protective enzymes was also carried out between the larvae with different levels of intoxication. It was found that the activities of protective enzyme of the seriously intoxicant larvae were higher than that of the lightly intoxicant larvae. This difference was more obvious in the group treated with CpTI transgenic poplar.

Increasing Accumulation Level of Foreign Protein in Transgenic Plants Through Protein Targeting

Targeting of the synthesized polypeptide in the cells is an important research field in modern cell biology. Cowpea trypsin inhibitor (cpti) gene has been modified and a fusion protein gene (sck) was produced by fusing a signal peptide sequence at cpti 5' end and an endoplasm reticulum (ER) retention signal peptide at cpti3' end respectively. The signal peptide can direct the newly synthesized polypeptide into ER, while ER retention signal can make the protein retained in the ER and its derivative protein body. ELISA test indicated that the accumulation level of foreign CpTI protein in sck transgenic tobacco (Nicotiana tabacum L.) was two times higher than cpti transgenic tobaccos and some individuals were four times higher. At the same time, sck transgenic tobacco has a high resistance to Lepidoptera pest due to the increased accumulation level of foreign CpTI protein. The strategy of foreign protein targeting can be used to increase the accumulation level of foreign protein in transgenic plants and can be widely applied to other related research field in plant genetic engineering.

Preliminary Study on the Separation of Lysozyme,Ovotransferrin and Ovalbumin from Egg White

[Objective] A study on separation process of lysozyme, ovotransferrin and ovalbumin from egg white. [Method] The proteins were separated by ammonium sul-fates and ion-exchange chromatography. Purity of the proteins was assayed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis （SDS-PAGE）. [Result] The results showed that the proteins were electrophoresis-pure. The specific activity of lysozyme was increased from 144.13 to 2 235 U/mg, and purification factor was 15-fold. Lysozyme recovery rate was estimated to be 15.76%. Bacteriostasis rate of ovotransferrin was 48.84%. [Conclusion] The procedure for separating lysozyme, ovotransferrin and ovalbumin from egg white was simple, fast, low-cost and suitable for industrilization.