As far as the enzymatic activity is concerned , on the one hand , strain yz - ii6 has higher d - hydantoinase activity and lower d - carbamoylase activity so as not to be suitable for one - step bioconversion of d - amino acids , on the other hand , the higher hydantoinase activity , engineered strain in particular , may be convenient to be as a biocatalyst to produce n - carbamyl d - amino acids which hard to find in the markets Yz - 6菌具有较高的海因酶活性，但n -氨甲酰基d -氨基酸酰氨水解酶活性很低，仍不适合用于d -型氨基酸生产工艺中的一步法转化。另一方面，含海因酶基因的人工菌株不但酶活性高，也排除了天然菌株转化生产d -型氨基酸过程中的一些副产物。
Objective : to clone and sequence the cdna encoding metalloproteinase from the venom of agkistrodon acutus from northen mountain area of guangxi province . methods : one step method was used to extract total rna from the venom of agkistrodon acutus found in northern mountain area of guangxi province . different kinds of cdna encoding metalloproteinase were amplified by one step method ( rt - pcr and pcr reactions occurred in the same tube ) using different primers 方法：从桂北五步蛇毒腺中抽提总rna ，利用不同的引物，采用一步法（ rt - pcr和pcr在同一管内进行）扩增出不同的dna条带，利用平端连接的方法将pcr扩增产物克隆至pgem - teasy载体，转化大肠杆菌jm109 ，挑选白色菌落提取质粒，用pcr对其进行鉴定，直接利用纯化pcr产物或提取阳性菌落质粒进行测序。
Water steam was used as oxidant , and the optimum water steam partial pressure is between 1 10 - 4 and 5 . 5 10 - 4 pa . under the optimum growth parameters , a ceo _ 2 seed layer with highly textured degree was successfully prepared . beside the one step process was experimented in this dissertation , the two step process was proposed and studied to further improve the quality of ceo _ 2 seed layer . in the two step process , about 15 nm thick of ce metal layer was deposited on metallic substrate at the first step , then water steam was introduced in the chamber , and the ceo _ 2 thin films were subsequently deposited with reactive sputtering in the 总结出沉积ceo _ 2薄膜的优化工艺条件,当沉积温度为720 - 850 、水蒸汽分压介于1 10 - 4 - 5 . 5 10 - 4pa之间、退火时间40min时,获得了织构程度良好的ceo _ 2种子层薄膜; 3 .由于一步法制备ceo _ 2种子层中水分压范围狭窄,工艺条件难以控制,并且退火延长了薄膜的制备时间,因此,本论文又采用了两步生长法沉积ceo _ 2种子层,即:先在ni - w基带上沉积一层约15nm的金属ce薄膜,再通入氧化气氛(水蒸汽) ,继续进行薄膜沉积。
The first one was called one step process or isothermal deposition and annealing process . in this process , the ceo _ 2 layers were formed at high temperature and oxidative atmosphere and then annealed at the same temperature . the relationship between the growth parameters and the textured degree of ceo _ 2 thin film was systematically studied , and the optimal growth parameters were summaried 采用等温退火法(或称“一步法” )沉积ceo _ 2 ,即:先在氧化性气氛下直接反应生长ceo _ 2薄膜,再在与沉积温度相同的温度下对薄膜进行退火处理,系统研究了沉积温度、退火时间、水蒸汽分压对薄膜c轴织构程度的影响。
In order to further investigate the role of axudl in human tumor carcinogenesis and the potential association between the axudl gene expression status and the stimulation of transforming growth factor beta in human cancers , the present study was performed in three aspects as follows : ( 1 ) cloning full length enconding region cdna of axudl and construction of eukaryotic vector that expression the fusion protein of axud1 and influenza virus hemagglutin ha epitope tag ; ( 2 ) exploring the time and dose effects of tgf - 1 on the expression - of axudl gene in hepg2 hepatoma cells and spc - a1 lung carcinomas cells , and studying the effects of overexpression of axud1 on the expression of cell cycle and apoptosis related protein in hepg2 hepatoma cells ; ( 3 ) construction and expression of human axudl in e . coli m15 . the following main results and conclusions can be obtained from the present study : 1 . the full length ecnoding region of human axudl cdna from human peripheral blood lymphocytes was successfully cloned using one step rt - pcr method , and constructed into a eukaryotic expression vector which can be expressed a ha - axud1 fusion protein with axud1 and influenza virus hemagglutin ha epitope tag . the recombinant plasmid was identified by polymerase chain reaction , restriction endonuclease maping and sequencing , this expression vector might be instrumental to further study the function of axud1 protein in tumor cells 为了进一步研究axud1在人类肿瘤发生中的作用及axud1基因的表达状况与tgf -介导的信号通路的关系，本实验研究分为三个部分： ( 1 ) axud1基因cdna全长编码区的克隆和ha表位标记的axud1基因表达载体的构建； ( 2 )探讨肝癌细胞hepg2和肺腺癌spc - a1细胞中tgf - 1诱导的axud1基因表达的时间、剂量效应以及诱导表达的可能机理，并研究axud1的过表达对细胞周期和细胞凋亡相关蛋白表达的影响； ( 3 ) axud1原核表达载体的构建及其在大肠杆菌中的表达。本实验的主要结果和结论如下： 1利用一步法rt - pcr成功地从人类外周血淋巴细胞中克隆出axud1基因编码区cdna ，并将其构建入真核表达载体中，编码的ha - axud1融合蛋白带有流感病毒凝血素ha的表位标记肽段。
Our efforts were taken to lay the foundation of further studies on cloning and function of new genes in fish . the construction and evalution of the a , gtlo cdna library of grass carp leukocytes are described . total rna was extracted from head - kidney leukocytes using single - step method of rna isolation by acid guanidinium thiocyanate - phenol - chloroform extraction 本文第一部分取病毒感染36小时后的草鱼头肾并分离白细胞，用改进的异硫氰酸胍一步法从其中提取总rna ，磁珠法分离纯化mrna ，电泳检测其质量。
