激光切割植物染色体的研究

文章探索了短脉冲激光与微细玻璃针相结合对黑麦染色体进行切割、分离的技术。该技术具有操作简便,节省经费,容易掌握,能减少染色体DNA的损伤,且可对整个细胞进行分离的优点,有利于促进染色体或染色体片段分离技术在进行DNA扩增,建立相应的DNA文库方面的普及和应用。     

黑麦B染色体显微切割和微克隆

显微分离出两条黑麦Ｂ染色体,经蛋白酶Ｋ处理,Ｓａｕ３ＡＩ酶切后,用ＬＡ－ＰＣＲ（ＬｉｎｋｅｒＡｄａｐｔｅｒＰＣＲ）方法进行扩增,得到０．２～２ｋｂ的ＤＮＡ片段．Ｓｏｕｔｈｅｒｎ杂交分析结果表明,此扩增产物来源于黑麦Ｂ染色体．将部分ＰＣＲ产物克隆到ｐＵＣ１９载体中,获得了５０００个克隆．为构建Ｂ染色体ＤＮＡ文库奠定了基础     

一种改进的染色体显微切割方法

本文介绍了利用普通光学显微镜进行染色体显微切割的方法．先使显微镜物镜焦点平面升高约7mm,将制在盖玻片上的染色体标本反扣在自制的玻璃台架上,从物镜下方进行染色体切割,这样就解决了在100×油浸物镜下不能切割染色体的困难,实现了在高放大倍数（放大率1000～1500×）条件下进行染色体显微切割的问题,大大提高了显微切割的精度和准确度．     

人类染色体中的微切割与微克隆技术

微切割、微克隆是分子生物学的一项新技术，它对于基因的标记与分离、DNA文库的重组及遗传图谱的绘制都发挥着重要作用．笔者介绍了这项技术在人类染色体中的应用情况．     

水稻染色体微切割及4号染色体酶解片段的扩增

通过细胞分裂的同步化,低渗,酶消化,瞬间固定和涂片等制片技术,快速制备水稻根尖体细胞染色体标本。该方法制备出的染色体标本,杂质少,染色体分散程度好,满足了染色体微切割用片的要求。染色体微切割和ＰＣＲ扩增技术,获得了水稻第４号染色体ＤＮＡ文库。     

激光显微切割技术在植物细胞和分子生物学研究中的应用进展

激光显微切割技术是20世纪90年代后期发展,由现代物理学和生物学相结合的精确取材技术.该技术可以在显微镜下利用微激光束直接从不同的组织(包括活体组织)快速切割、分离和纯化生物体的目标组织、细胞及其组分,用于细胞和分子生物学的研究.笔者结合开展激光显微切割技术研究遇到的问题,从制片技术的改进、基因表达、DNA和蛋白质分析等方面,对激光显徽切割技术在植物细胞和分子生物学研究中的应用进行了综述.     

激光切割顶针自动回避

在加工零件轮廓图形显示的基础上，采用区域分析的方法，通过逐段分析加工零件的ＮＣ代码，求出加工零件轮廓可避开顶针保护区的范围，以“最小偏移”原则修改ＮＣ代码，从而使零件轮廓避开顶针保护区，起到保证加工质量和保护顶针的作用．     

应用PCR—SSCP技术从显微切割的人染色体17q11—12探针池批量分离单拷贝片段的研究

将ＤＮＡ单链构象多态（ＳＳＣＰ）检测的原理，应用于“显微切割的人染色体１７ｑ１１—１２探针池”批量分离单拷贝片段，取得了很好效果。从筛选出的７４个次级单拷贝中有效地鉴定出３７个非同源的单拷贝片段。这比传统的仅限于长度比较而确认的非同源单拷贝片段（１２个）多出２５个．其中部分已经ＤＮＡ测序证实。结果提示：采用ＳＳＣＰ技术区分相似分子量单拷贝片段的同源性，可显著地提高从“显微切割探针池”分离和克隆非同源单拷贝片段的效率。本文还将一部分单拷贝片段作为探讨，与人基因组ＤＮＡ的限制性片段作Ｓｏｕｔｈｅｒｎ印迹杂交，结果均只显示单一杂交带，说明单拷贝ＤＮＡ片段的结论是可靠的。     

Chromosome microdissection by laser microbeam, chromosomal fragment isolation and amplificationin vitro in barley (Hordeum vulgare L.)

A method for microdissection, isolation and amplification of plant chromosomal fragments using laser microbeam and a glass microneedle was established. Firstly, 7H chromosome of barley (Hordeum vulgare L.) was dissected by Nd: YAG laserbeam with suitable parameters and the fragment comprising a satellite was isolated with a glass microneedle which was fixed on a micromanipulator. Then, the chromosomal fragment DNA was amplified by LA_PCR (linker adaptor PCR) for two rounds. The size of the DNA fragments of PCR products varied from 500-3 000 bp and the PCR products originated from the genome of barley were verified by Southern hybridization. Compared with previous reports, there are some advantages in this research. The performance is easier, the dissection is more precise and the cost is low. It also permits efficient amplification with only one single chromosome fragment. Laser microbeam_glass microneedle method may be useful in the microdissection of special chromosome regions, especially in plants with middle or small chromosomes.     

Microdissection and amplification of the chromosome arm 5S in a rice telo-tetrasomic

The rice aneuploids with telochromosomes are ideal genetic stocks for chromosome arm identification and microdissection. A rice telo-tetrasomic line with two extra short arms of chromosome 5 (5S) was used in the present study. The amrs of 5S were microdissected from the prometaphase cells in mitosis and amplified with linker adaptor PCR (LA-PCR). The amplified fmgments, ranging from 200–3000 bp, were confirmed to be from the rice chromosome arm 5S by the rice STS and miamatellite markers.     

Microdissection of chromosome 7B of common wheat and cloning of low-copy specific DNA sequences

The 7B chromosome of common wheat was microdissected from pollen mother cells of the 7B monosomic line of common wheat cv. Chinese Spring (CS). After proteinase K and DNA topoisomerase Ⅰ treatments, the isolated chromosomes were subjected to 1-3 rounds of DOP-PCR amplification, which produced continuous DNA fragments ranging from 150 to 700 bp. Ge-nomic Southern hybridization confirmed that the PCR products were originated from the wheat genome. Cloning of portion ( > 200 bp) of the 3rd round DOP-PCR products (50 μL) could generate about 20 000 recombinant clones. Characterization of 50 randomly chosen clones indicated that 21 clones produced discrete PCR products with the size of 240-600 bp. Dot-blot hybridization showed that among the 21 clones, 11 ( ～ 55%) were of low-copy nature while 10 ( ～ 45%) were repetitive. Southern hybridization with the complete set of the CS "nullisomic-tetrasomic (NT)" lines demonstrated that all the 6 low-copy clones were specific to either chromosome 7B or the 7th homoeologous group, whereas the 3 arbitrarily chosen repetitive clones were non-specific, disperse sequences.     

葎草单染色体的显微分离及DOP-PCR技术体系建立

葎草是具有XX/XY1Y2性染色体系统的雌雄异株植物,是研究植物性染色体演化的模式材料之一.利用染色体显微分离技术从葎草根尖有丝分裂中期分裂相中将单条染色体进行了显微分离及DOP-PCR(Degenerate oligonucleotide primer-PCR)扩增,并构建了单染色体DOP-PCR扩增产物的荧光探针,对葎草根尖有丝分裂中期分裂相染色体进行了荧光原位杂交,其结果表明荧光信号分布在所有的染色体上,表明所建立的技术体系能够成功分离葎草单染色体并进行DNA扩增.本研究结果为进一步进行葎草X,Y染色体的细胞及分子生物学研究提供了技术支持.     

基于单片机的自动控制系统的数字化改造

以双闭环直流脉宽调速为例,介绍了模拟控制系统和数字控制系统的实现方法,给出了具体的实现电路,对其它模拟系统的数字化具有一定的参考价值。     

植物激素对蚕豆胚轴愈伤组织染色体加倍和减倍的影响

植物激素能引起蚕豆愈伤组织染色体发生加倍和减倍，特别是ＮＡＡ５ｍｇ／Ｌ，ＫＴ０．５ｍｇ／Ｌ能获得较多的单倍体，同时ＮＡＡ在０～１ｍｇ／Ｌ之间，ＫＴ在２～５ｍｇ／Ｌ之间能获得较多的四倍体．通过对加倍和减倍的分析，推出加倍率与单倍体和四倍体频率的关系．     

罗氏沼虾的染色体研究

从罗氏沼虾处于囊胚期和原肠期的胚胎细胞中提取中期染色体进行细胞遗传学研究。染色体制片分析表明，罗氏沼虾二倍体染色体数目2N=118，但是也有一些细胞具有100～109条染色体。以外，还有一些细胞具有与单倍体染色体数相近的染色体数。