豌豆“怀孕”——F2种子种皮、子叶性状教学的突破

1例题分析 例题：豌豆灰种皮对白种皮为显性,黄子叶对绿子叶为显性。每对性状的杂合子（F1）自交后代均出现3：1的性状分离比。请问以上种皮颜色的分离比和子叶颜色的分离比分别来自于对以下哪代植株所结种子的统计？（ ）     

转抗真菌病基因烟草后代遗传分析

利用载体pBLGC 通过发根农杆菌（Agrobacterium rhizogines）叶盘法转化烟草。对烟草转基因植株后代从DNA水平、RNA水平以及几丁质酶基因和β－1,3葡聚糖酶基因表达效率方面进行系统的研究。对探讨转基因植株后代遗传规律具有重要的意义。     

辐照外源DNA导入番茄两种同工酶的研究

采用聚丙烯酰胺梯度凝胶电泳对辐照外源DNA导入番茄的受体后代进行过氧化物酶和细胞色素氧化酶同工酶研究,并对供体叶形在受体后代中的表达和后代株高进行分析。研究表明,多数受体后代与受体对照的两种同工酶图谱差异较大而与供体对照的两种同工酶图谱较为相似,这与它们在外观性状（叶形和株高）上的差异是吻合的。受体后代与体供在同工酶图谱上的相似性以及供体外观性状在受体后代的表达表明了外源DNA已经导入受体,并得到整合表达,说明辐照外源DNA导入技术是改良番茄品种,丰富育种材料的一条有效途径,在番茄选育新品种方面有良好的应用前景。     

快中子辐照结合组织培养培育花生新品种宇花7号

为开拓新的花生育种方法,对辐照诱变结合组织培养创造花生新种质、培育新品种进行了研究。以我国北方地区主栽花生品种鲁花11号成熟种子为试材,经快中子辐照处理后取种子胚小叶进行组织培养,通过胚胎发生途径获得再生苗。再生苗经嫁接驯化后移栽田间,83个单株获得种子。后代按系谱法进行选育,从83个再生植株后代中获得了107份突变体,分别在主茎高、分枝数、荚果形状和大小、种皮颜色、内种皮颜色、含油率、蛋白含量等性状上发生了明显变异。从突变体后代中选育出了低油早熟耐涝大花生新品种宇花7号,其产量比亲本鲁花11号增产14%以上;其含油率(47.0%)比鲁花11号低5.1个百分点。宇花7号2016年参加辽宁省新品种登记试验,比对照品种白沙1017平均增产13.8%。2018年通过了国家非主要农作物品种登记,登记号为"GPD花生(2018) 370105"。研究结果说明,辐照结合组织培养是创造花生新种质、培育新品种的有效方法。     

2003年国际生物学奥林匹克竞赛实验考试试题(5)

第四实验室:遗传学实验实验1菜豆种皮颜色遗传方式的遗传分析完成本题的时间是25min材料与设备亲代种子(P1)——(样品No.1)亲代种子(P2)——(样品No.2)杂交种子(F1)——(样品No.3)分析用品系种子(La)——(样品No4)Fa带种子——(样品No.5)盛种子的盘子2个白纸1张菜豆种皮颜色由一系列基因控制,这些基因分别控制色素的合成,种皮颜色的分布,以及增色、淡化或者以某种方式改变颜色的修饰基因。在预实验中,两种种皮颜色不同菜豆(P1和P2)的杂交育种实验。用杂交子1代的种子进行栽培实验,长成的杂合个体(F1)得F1表型种子。下一阶段实验用子一…     

玉米DNA导入水稻的RAPD分子验证

对高光效C4植物玉米DNA为供体、通过花粉管通道技术导入水稻所获得的水稻后代进行了RAPD分析.结果表明:从60个引物中找到12个引物检测出DNA的多态性,证明外源DNA导入受体后引起后代基因组的显著变异.     

拟南芥种皮色素形成突变体的筛选与表型鉴定

类黄酮在植物应答各种环境胁迫和种皮发育调控中起着重要作用。通过甲基磺酸乙酯（EMS）诱变筛选获得1个透明种皮突变体,与野生型拟南芥（Arabidopsis thaliana）(Col-0)相比,突变体成熟的种子颜色为黄色,其表型性状由隐性单基因控制。利用图位克隆和精细定位技术将突变基因定位于5号染色体MAH20的BAC上,是TT4(At5G13930)基因的第1 299位碱基C突变为T,使得第324位氨基酸甘氨酸突变为谷氨酸。TT4(transparent testa 4)编码1个类黄酮合成的结构基因查尔酮合酶（CHS）,突变后种皮透明,种子颜色为黄色,突变体命名为tt4-1。利用功能回补突变体恢复褐色种皮表型,进一步证明了TT4在调节种皮颜色发育过程的重要作用。启动子偶联GUS基因组织表达分析显示TT4基因在植株幼苗的根、茎、叶和花中均有表达,生理表型分析结果显示与野生型相比,突变体tt4-1种子萌发早,幼苗主根短、侧根和根毛较多,成苗叶片气孔开度大和失水率高等特性。该研究将为进一步阐述TT4基因功能奠定理论依据。     

嫁接导致的绿豆可遗传性变异及其在绿豆育种中的应用

植物嫁接可以导致接穗的后代中有可遗传性变异的发生.我们将绿豆(Vigna radiata (L.) Wilczek)的幼苗嫁接在红薯(Ipomoea batatas (L.) Lam.)的茎上, 维护其生长至结实.将收获的绿豆种子连续几代播种于普通环境时, 其后代中出现了明显的遗传变异.这些变异在未经嫁接的绿豆接穗品系中并不出现.为了研究这种嫁接诱导变异现象可能的机理, 我们对变异品系的细胞质和细胞核DNA进行了分析.结果显示,在原绿豆和变异品系之间未发现细胞质DNA的RFLP(限制性酶切片段多态性)差异.而细胞核DNA却发生了高频率的序列重组.同时,本研究没有发现砧木与接穗间基因转移的迹象.根据以上事实,我们推测远缘嫁接变异很有可能是嫁接生长逆境诱导的抗逆变异.     

遗传教学应注意种子颜色与种皮颜色的区分

孟德尔在杂交实验中曾用过豌豆种子颜色这一相对性状,种子的颜色是指从种皮透出的子叶的颜色。但日常提到的种子颜色,大部分都不是真正种子颜色而是种皮的颜色,由于种皮是属于种子的一部分,所以一般情况这样提也还可以,但在讨论遗传问题时就应从发生角度予以区分,否则就会出现实验与理论的出入。把种子颜色定为子叶的颜色,就说明种子颜色决定了该种子自身的基因型。种子和果皮分别是由珠被和子房壁发育来的,因而它们实质上是母体分离出来的一部分,其基因型与母体的体细胞是相同的,杂交不可能改变这部分的基因型,而只能改变由受精的卵细胞和极核发育来的胚和胚乳细胞之基因型。所以杂交当代结     

DNA复制的忠实性

染色体DNA携带着生物的遗传信息,生物依赖传递核酸以繁殖后代.产生与亲代等同的子代细胞需要在DNA复制时,子代DNA的碱基序列忠实于亲代DNA的碱基序列. DNA复制主要由DNA聚合酶催化.聚     

氯化钠密度梯度离心法制备用于显微注射的外源DNA片段

DNA显微注射是生产转基因动物最可靠和最常使用的一种方法,外源DNA的纯度对显微注射的成功起着至关重要的作用。本文介绍用氯化钠密度梯度离心的方法制备用于显微注射的外源DNA片段。与传统的琼脂糖凝胶回收的方法相比较,用此方法制备的外源DNA片段对小鼠受精卵进行显微注射后,受卵体母鼠的胚胎存活率,以及子代小鼠的外源基因整合率均有明显的提高。这一方法可为进一步提高转基因动物的成功率,提供方法学上的参考。 Abstract:DNA microinjection is the most popular and reliable method of producing transgenic animals.The purity of foreign DNA plays an important role for the success of microinjection.In this study,we introduced the use of sodium chloride step gradients in fractionating foreign DNA fragment for microinjection.The data demonstrated that,compared with the conventional agarose gel extraction method,NaCl purification scheme of toreign DNA could improve the treated embryo survival and foreign DNA intergration rate markedly.     

On the fate of orally ingested foreign DNA in mice: chromosomal association and placental transmission to the fetus

We have previously shown that, when administered orally to mice, bacteriophage M13 DNA, as a paradigm foreign DNA without homology to the mouse genome, can persist in fragmented form in the gastrointestinal tract, penetrate the intestinal wall, and reach the nuclei of leukocytes, spleen and liver cells. Similar results were obtained when a plasmid containing the gene for the green fluorescent protein (pEGFP-C1) was fed to mice. In spleen, the foreign DNA was detected in covalent linkage to DNA with a high degree of homology to mouse genes, perhaps pseudogenes, or to authentic E. coli DNA. We have now extended these studies to the offspring of mice that were fed regularly during pregnancy with a daily dose of 50 g of M13 or pEGFP-C1 DNA. Using the polymerase chain reaction (PCR) or the fluorescent in situ hybridization (FISH) method, foreign DNA, orally ingested by pregnant mice, can be discovered in various organs of fetuses and of newborn animals. The M13 DNA fragments have a length of about 830 bp. In various organs of the mouse fetus, clusters of cells contain foreign DNA as revealed by FISH. The foreign DNA is invariably located in the nuclei. We have never found all cells of the fetus to be transgenic for the foreign DNA. This distribution pattern argues for a transplacental pathway rather than for germline transmission which might be expected only after long-time feeding regimens. In rare cells of three different fetuses, whose mothers have been fed with M13 DNA during gestation, the foreign DNA was detected by FISH in association with both chromatids. Is maternally ingested foreign DNA a potential mutagen for the developing fetus? Received: 15 April 1998 / Accepted: 15 June 1998     

用Digoxigenin标记DNA探针作基因转移山羊的整合检测

对采用外源基因原核显微注射法生产的基因转移山羊,用一种新的DNA非同位素标记法即地高辛配基标记DNA探针作外源基因整合检测.结果表明:地高辛标记核酸探针能检测出基因转移动物基因组中整合的外源基因.     

电脉冲作用将外源基因导入稀有(鱼句)鲫精子的研究

将稀有鲫 (Gobiocyprisrarus)精子与重组质粒pCAhLFc线性DNA混合温育 ,经电脉冲处理后与卵子受精 ,孵化出苗。从鱼苗中提取DNA ,经PCR检测 ,2 5 .5 %～ 6 6 .7%鱼苗带有外源基因。在显微镜下观察经电脉冲处理过的精子 ,发现其活力有不同程度下降 ,受精率也有不同程度下降 ,说明不同的电脉冲条件对精子有不同程度的损害作用。精子与外源DNA混合温育 ,经电脉冲处理后 ,用DNA外切酶消化后 ,提取精子DNA ,经PCR检测 ,仍有阳性电泳带 ,证明电脉冲可以促使稀有鲫精子摄入外源基因     

Co-injection of restriction enzyme with foreign DNA into the pronucleus for elevating production efficiencies of transgenic animals

The microinjection method for production of transgenic farm animals requires specialized techniques and results in intolerably low production efficiencies. We investigated whether or not co-injection of foreign DNA constructs with restriction endonuclease into the pronucleus of mouse zygotes would improve the integration frequencies of foreign DNA into the host genome. Two kinds of DNA constructs that have no EcoRI site in their sequences were used for co-microinjection. With reference to the results of experiments in which EcoRI alone was injected at various amounts varying from 10(-9) to 10(-5) U/nucleus, the amount of 5x10(-8) U/nucleus that showed survival rate of 60.6% was used for the co-injection with DNA. Successful transgenesis of co-injected embryos was identified by DpnI-Bal31 digestion method for single embryos and by PCR method for pups born, respectively. The overall efficiency for the integration of foreign DNA in single embryos and live-born pups obtained by the co-injection procedures were 17.9% compared with 9.1% obtained by the injection of DNA alone. The results suggest that co-injection of foreign genes with restriction enzyme may elevate the integration rate of foreign genes into host genomes.     

Stable integration of foreign DNA into the chromosome of the cyanobacterium Synechococcus R2

The blue-green alga, Synechococcus R2, is transformed to antibiotic resistance by chimeric DNA molecules consisting of Synechococcus R2 chromosomal DNA linked to antibiotic-resistance genes from Escherichia coli. Chimeric DNA integrates into the Synechococcus R2 chromosome by homologous recombination. The efficiency of transformation, as well as the stability of integrated foreign DNA, depends on the position of the foreign genes relative to Synechococcus R2 DNA in the chimeric molecule. When the Synechococcus R2 DNA fragment is interrupted by foreign DNA, integration occurs through replacement of chromosomal DNA by homologous chimeric DNA containing the foreign insert; transformation is efficient and the foreign gene is stable. Mutagenesis in some cases attends integration, depending on the site of insertion. Foreign DNA linked to the ends of Synechococcus R2 DNA in a circular molecule, however, integrates less efficiently. Integration results in duplicate copies of Synechococcus R2 DNA flanking the foreign gene and the foreign DNA is unstable. Transformation in Synechococcus R2 can be exploited to modify precisely and extensively the genome of this photosynthetic microorganism.     

A new concept in (adenoviral) oncogenesis: integration of foreign DNA and its consequences

A new concept for viral oncogenesis is presented which is based on experimental work on the chromosomal integration of adenovirus DNA into mammalian genomes. The mechanism of adenovirus DNA integration is akin to non-sequence-specific insertional recombination in which patch homologies between the recombination partners are frequently observed. This reaction has been imitated in a cell-free system by using nuclear extracts from hamster cells and partly purified fractions derived from them. As a consequence of foreign DNA insertion into the mammalian genome, the foreign DNA is extensively de novo methylated in specific patterns, presumably as part of a mammalian host cell defense mechanism against inserted foreign DNA which can be permanently silenced in this way. A further corollary of foreign (adenovirus or bacteriophage λ) DNA integration is seen in extensive changes in cellular DNA methylation patterns at sites far remote from the locus of insertional recombination. Repetitive cellular, retrotransposon-like sequences are particularly, but not exclusively, prone to these increases in DNA methylation. It is conceivable that these changes in DNA methylation are a reflection of a profound overall reorganization process in the affected genomes. Could these alterations significantly contribute to the transformation events during viral or other types of oncogenesis? These sequelae of foreign DNA integration into established mammalian genomes will have to be critically considered when interpreting results obtained with transgenic, knock-out, and knock-in animals and when devising schemes for human somatic gene therapy.The interpretation of de novo methylation as a cellular defense mechanism has prompted investigations on the fate of food-ingested foreign DNA. The gastrointestinal (GI) tract provides a large surface for the entry of foreign DNA into any organism. As a tracer molecule, bacteriophage M13 DNA has been fed to mice. Fragments of this DNA can be found in small amounts (about 1 % of the administered DNA) in all parts of the intestinal tract and in the feces. Furthermore, M13 DNA can be traced in the columnar epithelia of the intestine, in Peyer's plaque leukocytes, in peripheral white blood cells, in spleen, and liver. Authentic M13 DNA has been recloned from total spleen DNA. If integrated, this DNA might elicit some of the described consequences of foreign DNA insertion into the mammalian genome. Food-ingested DNA will likely infiltrate the organism more frequently than viral DNA.     

Instability of hybrid plasmids containing Drosophila melanogaster DNA in rec+ and rec- Escherichia coli K-12 strains

The stability of hybrid plasmids, constructed on the basis of vector pCV20(AprTcr) and containing HindIII fragments of Drosophila melanogaster DNA (pDm6, pDm9) and PstI fragments of D. melanogaster DNA (pDm39, pDm187, pDm189) was studied. After the transformation of E. coli HB101 recA and Escherichia coli 802 rec+ and selection to Tcr (pDm6, pDm9), or to Apr (pDm39, pDm189, pDm187) 0.04--9% of clones with reduced resistance to Tc or Ap was detected. The hybrid plasmids are more stable in rec-, but not in rec+ strain, the stability depends of the nature of cloned DNA, and on the site of vector DNA in which foreign genes are cloned. Restriction endonuclease analysis revealed that all plasmids of the clones with reduced Tcr or Apr lost the inserted DNA and the excision of foreign DNA occurred precisely in the sites of cloning. We suggest that the genome of the hybrid plasmid in the region of foreign insertion has a conformation which allows the bringing together the ends of cloned DNA with the following excision of the foreign genes.