( 2 ) average insert size of the library is 118 kb evaluated from analysis of 452 bacs randomly picked from library . so the genome coverage of the library is 13 . 34 and the possibility to find a single - copy gene in the library is 99 . 984 % ( 2 )对随机挑选的452个bac克隆的插入片段进行鉴定估计文库的平均插入为118kb ，文库的基因组覆盖率为13 . 34倍，预计从文库中筛选到单拷贝基因的机率为99 . 984 。
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。
The cyp51 genes cloned from both isolates of imazalil - resistant and imazalil - sensitive of p . digitatum were cloned into the pcb 1004 expression vectors , respectively . the direction and number of inserted copy were confirmed by restriction enzyme digestion 本研究构建了指状青霉对抑霉唑敏感和抗性的两个cyp51基因的pcb1004表达载体，并对基因的插入方向和拷贝数进行了分析，证明了两个载体连接的基因均为正向连接和单拷贝插入。
Results showed that : ( 1 ) in general , the segregation ratios of target gene in some tl lines are conformed to 3 : 1 , however , some are not possibly due to gene silence or missing in the self - pollinated progeny ; ( 2 ) two homozygous plants were identified from 10 putative transgenic plants 主要结果如下： ( 1 )我们对外源基因在番茄体内的遗传规律研究表明，在大多数情况下，单拷贝插入外源基因将导致转基因后代按孟德尔单显性基因3 1规律分离，但是部分转化系比例偏低，可能发生了基因丢失或者沉默。
1 . because the taxonomic division is rather complex and has been much disputed and revised , in this part , we will review the classification and phylogeny of families , subfamilies and tribes of anseriformes based on morphology , ethology , osteology , mitochondrial and nuclear dna restriction fragment length polymorphism , single - copy nuclear dna hybridization and the sequences of mitochondrial gene analysis referring to the different definition , classification and phylogenetic relationships of the families , subfamilies and tribes of anseriformes . the controversial questions and deficiency in the systematic studies of anseriformes were pointed out 具体包括以下几个部分： 1 、针对雁形目鸟类异常复杂的分类状况及分类上存在的争议，根据雁形目鸟类的形态学、行为学、骨骼学、角蛋白、线粒体与核dna酶切片段长度多态、单拷贝核dna - dna杂交及线粒体基因dna序列分析等方面的研究，对雁形目鸟类分类中科、亚科和族的划分及其相互间的系统发生关系进行综述，分析系统学研究中存在的不足，提出了雁形目鸟类分类中急需解决的问题。
Objective : construct high - level expression system of echistatin in e . coli methods : obtain amino - acid sequence of echistatin from genebank database . considering the bias of usage of 61 available aminoacid codons in e . coli , design the coding sequence of echistatin , synthesize the dna sequence chemically , get single copy coding gene and repeated two copy coding gene of echistatin . insert the sequence into expression vector pbv220 , and more , we construct fusion expression clone of echistatin with pcr , identify the recombinant vector by dna sequencing 目的构建蛇毒锯鳞蝰素( echistatin )的原核高效表达体系方法由genebank数据库检索蛇毒锯鳞蝰素( echistatin )的氨基酸序列，结合大肠杆菌蛋白质合成体系对氨基酸密码子使用的偏爱性，设计了echistatin编码基因，体外人工合成编码基因dna片段，通过适当的限制性内切酶位点插入表达载体pbv220 ，分别构建了echistatin的单拷贝表达克隆、双拷贝串联表达克隆；进一步通过pcr技术构建echistatin的融合表达基因克隆。
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 （ 。
Then , we transformed those two genes into tomato and tobacco plants via agrobacterium tumefaciens . after verified by antibiotic resistance , reporter gene examination , southern blot detection , and genetic segregation analysis , we obtained 3 and 7 transgenic tobacco plants with one copy of rbcs 3a - gus or rbcs 3c - gus gene , respectively . further , we established two suspension - cultured cell lines using above mentioned two kinds of transgenic tobacco plants 对得到的再生烟草植株分别进行了报告基因表达水平检测、 southern - blot鉴定以及t _ 0代转基因种子遗传分离规律分析，分别得到了单拷贝的稳定表达番茄rbcs3a - gusa 、 rbcs3c - gusa和35s - gusa基因的转基因烟草3株、 7株和1株，同时还得到单拷贝的只转化gusa基因的阴性对照转基因烟草3株。
Results : designed and synthesized the coding gene of echistatin . and constructe its single copy expression vector , identify the vector by dna sequencing . but it could not express echistatin in e . coli . designed and synthesized the modified coding dna of echistatin , and constructe its two copy expression vector , identify the vector by dna sequencing , but it could not express echistatin in e . coli 结果设计并合成了echistatin编码基因dna序列，构建了echistatin单拷贝表达载体，经dna测序鉴定正确后， sds - page及western - blotting结果表明： echistatin在上述菌株中未获得高效表达。
The time of silencing , and the percentage and state of silenced plants were recorded , and it was noted that the efficiency of induction of homologous dependent gene silencing ( hdgs ) by the gfp tnos vector was markedly higher than mat resulting from other vectors with intact structure 根据发生沉默的时间早晚、发生沉默的植株比例以及程度的观察，发现gfp - tnos载体诱发同源基因沉默的效率明显高于含有完整表达单元的同类载体；低拷贝载体诱发同源基因沉默的效率明显高于含有相同表达单元的单拷贝载体。
