| 1. | Gene duplication is defined as segment of dna duplicate one or more copies , it is believed to be a major mechanism for the generation of evolutionary novelty
摘要基因倍增指基因组中含有基因的dna片段复制出一个或更多拷贝的过程，是进化出新物种的主要原因。 |
| 2. | The whole gene was added every six hours instead of all in one time . in proper time , they were mixed and continued to ligate to multi - copy in the same direction
通过控制基因和接头加入的量及次序，可得到两侧有ecor和sal酶切位点的同向串连的多拷贝基因。 |
| 3. | Multicopy clones were screened from vary levels of g418 - ypd plate and cultured and induced with methanol to secrete interesting peptide , which was identified by sds - page and western blotting
培养多拷贝插入的酵母细胞，用甲醇诱导目的蛋白的分泌， sds - page和westernblotting鉴定表达蛋白。 |
| 4. | After linerization , the recombinant plasmid was transformed into pic hia pastoris by electroporation , which then were cultured in md plate free of histidine , from which the positive colones were propagated
重组质粒线性化后，用电击法将重组质粒转化入巴氏毕赤酵母，在缺组氨酸的md板上筛选阳性菌落，然后用不同浓度的g418 ? ypd板筛选多拷贝插入单菌落。 |
| 5. | 2 . southern blotting analysis indicated that sod2 gene had been integrated into all of the transgenic plants ' genomes . the copy number of sod2 in transgenic plants was mostly 1 to 5 by southern analysis
Southern结果表明，野生型植株(对照)没有明显的杂交信号，转基因株系有强阳性信号，转基因植株中插入外源基因的拷贝数不同，既有单拷贝的，又有两个拷贝及多拷贝的，拷贝数从1 - 5不等。 |
| 6. | Based on the blast analysis and other studies , osddl mutant was a multi - copy - insertion mutant , and one of the insertion sites was in an nptii like transposase gene , whereas osdd2 , a single - copy - insertion mutant was disrupted at a gene encoded wrky transcript factor
Osdd1突变体可能是多拷贝插入，其中一个插入到nptii转座酶类似基因。 osdd2突变体为单拷贝、插入到一个wrky类转录因子基因5翻译起始区附近。 |
| 7. | Multicopy integrants were screened with g418 from pichia pastoris which contains recombinant plasmid , and induced with methanol to secrete interesting peptide . the supernate of pichia pastoris culture was analysed by sds - page and western blotting . a reactive band , which the apparent molecular weight is 36kd , can be detected with sheep anti - hcmv polyclonal antibodies
重组质粒转化巴氏毕赤酵母， g418筛选出多拷贝插入的单克隆，甲醇诱导多拷贝插入的单克隆酵母细胞分泌目的蛋白，培养液上清经sds - page电泳分析，在蛋白质印迹中检测到培养液上清有一表观分子量为36kd ，能与羊抗hcmv多克隆抗体发生发应的条带。 |
| 8. | Multi - copies insertion transformants were screened on g418 plates . the recombinant protein was proved to have biological activity of hydrolyzing n - carbamoylphenylalanine into phenylalanine through enzyme activity assay . the n - carbamoylase activity of recombinant was 2 . 26 and 2 . 15 times higher than that of arthrobacter bt801 and dh5a / puc18 - 169
将hyucdna片段连接到真核表达载体ppic3 . 5k上，经bgl酶切线性化，通过peg法转化导入毕赤酵母gs115感受态细胞，利用g418抗性筛选得到12个插入多拷贝目的基因的转化子。 |
| 9. | Methods : 1 . expression and purification of recombinant human ctla4 extracellular domain : a 400bp dna fragment of ctla4 extracellular domain was obtained from the pctla4 / ig plasmid with genetical technique . then , this dna fragment was inserted into ppic9 plasmid to construct the single copy plasmid of yeast expression system ( ppic9 - ctla4125 ) . furthermore , the multi - copy plasmid ( ppic9k - ctla4125 ) was constructed
重组人ctla4胞外区蛋白在酵母gs115中的表达和纯化：首先通过基因操作技术，从pctla4八g质粒中扩增400hp的ctla4胞外区片段；将ctla4胞外区片段插人ppicg质粒中获取单拷贝酵母表达质粒ppicg ctla4125并进一步构建多拷贝酵母表达质粒ppicgk （ 。 |
