中国普通野生稻核糖体RNA基因限制性片段长度多态性

对９８份普通野生稻、亚洲栽培稻及稻进行了核糖体ＲＮＡ基因间间隔区的限制性片段长度多态性分析。共发现３０种长度变异类型,组成４５种表现型。广西普通野生稻的ｒＤＮＡ间隔序列长度多样性最丰富,２４份材料中长度变异类型,     

中国普通野生稻核糖体RNA墓因限制性片段长度多态性

对98份普通野生稻、亚洲栽培稻及稻进行了核糖体RNA基因间间隔区的限制性片段长度多态性（RFLP）分析。共发现30种长度变异类型,组成45种表现型。广西普通野生稻的rDNA间隔序列长度多样性最丰富,24份材料中检测到25种长度变异类型,共组成22种表现型。rDNA间隔区长度变异类型的地理分布与纬度相关,高纬度来源的普通野生稻的rDNA间隔区长度较短,与粳稻的长度变异类型相似;低纬度来源的材料rDNA间隔长度趋长,与籼稻的长度变异类型相似;稻的长度变异类型与粳稻相似。提出了华南是野生稻变异中心的观点,并指出对我国野生稻资源进行保护的重要性。     

对TMV不同抗性番茄品种叶绿体DNA限制性差异片段的克隆

对ＴＭＶ不同抗性番茄品种ｃｔ－ＤＮＡ经限制性内酶切后,将所获得的差异片段Ｂａｍ６克隆到质粒ｐＢｌｕｅｓｃｒｉｐｔⅡＳＫ中,经筛选、菌落原位杂交、斑点杂交及电泳鉴定,证明重组体为抗性品种Ｂａｍ６差异片段克隆,对探讨番茄ｃｔ－ＤＮＡ与抗ＴＭＶ的关系有重要意义。     

萝卜叶绿体DNA花粉育性特异片段的定位

本实验采用萝卜细胞质雄性不育系401A及其同核保持系401B为材料,利用BamH I、EcoR I、BglI和HindⅡ四种内切酶酶解其叶绿体：DNA。除HindⅡ外,其余三种内切酶电泳图谱中均显示两系之间的明显差异。将BamH I的5个差异片段分别与载体pBR322进行体外连接,获得4种重组体克隆。利用3种膜蛋白质基因探针,分别与两系的叶绿体DNA杂交,杂交均未出现在差异片段所在部位,这说明,两系之间的差别可能与这3种基因无关。将重组体质粒分别与P700叶绿素a脱辅基蛋白质基因探针及反向重复区内rRNA基因区探针杂交,结果表明,有3种重组体质粒所携带的差异片段与rRNA基因探针在杂交中显示出明显的阳性反应。也就是说,这3个差异片段均位于rRNA基因所在的叶绿体基因组的反向重复区中。     

小克银汉霉核糖体DNA PCR扩增产物的限制性片段长度多型性

应用一对寡核苷酸引物ITSl与ITS4对核DNA G+C百分数小于30％的小克银汉霉(Cunninghamella)属的核糖体DNA(rDNA)内转录间区(ITS)进行了扩增。测试的属于10个种和变种的22株菌都得到了扩增产物。在同属不同种之间扩增的ITS片段长度有巨大差别。据此可分为三组。这三组所含分类群数及其DNA长度分别为：第一组4种1变种,小于 764碱基对(bp);第二组2种,765～824 bp;第三组2种l变种,大于825 bp。所研究的许多种中,特别是第三组,单凭其PCR产物的长度就能区分开来。但在第一、二组就需借助限制性内切酶的分析才能予以区分。我们选用了Rsa I,Tru 9 I,和Hinf I 4种限制性内切酶,对所有扩增产物进行了限制性片段长度多型性(RFLPs)的分析。在雅致小克银汉霉(C elegans),巴西玉蕊小克银汉霉(C.bertholletiae)、和布拉氏霉(C. blakesleeana )各种的限制性酶切图谱,种内非常一致而种与种之间有差别。反之,在某些种其限制性酶切图谱不仅种间互不相同,在种内不同株之间也出现1～3种酶切图型的差异。在这种情况下,只能综合各种资料才能将它们区分开来。本研究肯定了PCR-RFLP在区分小克银汉霉种和变种上的意义,并发现种内个体的差异。这在我们后来所进行的序列分析研究中得到了进一步的证实。     

植物叶绿体4.5S核糖体rRNA作为分子杂交探针的研究

我们提取纯化了芹菜,菠菜和蕃茄叶绿体核糖体4.5SRNA(4.5SrRNA)并在其5’端标记~(32)P,作为探针与菠菜,蕃茄和芹菜叶绿体DNA(ctDNA)进行分子杂交。结果不仅证明4.5SrRNA可作为公用分子杂交探针,而且也说明不同植物4.5SrRNA序列有相当大的同源性。     

萝卜叶绿体DNA花粉育性有关的特异片段的部分序列组成

本实验序列分析并精确定位了萝卜(Raphanus sativus)叶绿体DNA花粉育性片段B_(21)的部分序列(ZL1)。结果表明,ZL1片段长474bp,其碱基组成是AT丰富的。与烟草全序列的比较分析发现,该片段位于烟草全序列中反向重复区IR_A的142330到142803、IR_B的100199到99726区段,其核苷酸序列与相应的烟草序列比较有96.6%的同源性。该片段具有rps12—rps7操纵子的一部分结构,分别编码核糖体小亚基蛋白S12的C-末端的7个氨基酸残基和S7的N-末端的93个氨基酸残基。两者的氨基酸序列与烟草、玉米、地钱、眼虫藻,大肠杆菌及蓝细菌相应序列的同源性分别为71.4—100%及40—95.5%。 上述结果意味着叶绿体核糖体蛋白质可能和细胞质雄性不育性存在某种联系。     

活化石植物－－苏铁树叶绿体DNA分子中的核糖核苷酸

随着分子生物学的深入发展,科学工作者对脱氧核糖核酸(DNA)的研究也更加深入。近年来发现一些植物叶绿体DNA(cpDNA)、动物线粒体DNA(mtDNA)分子中含有少数核糖核苷酸。其中对豌豆,菠菜和莴苣叶绿体DNA分子中的核糖核苷酸研究较多,而且比较详细。研究这个问题,使用的方法是碱(KOH)或核糖核酸酶(RNase)处理cpDNA。如果DNA分     

Preparation of Chloroplast DNA from Barley and Lettuce and Comparison of Restriction Fragments

Chloroplast DNA from three barley cultivars and from one lettuce cultivar was prepared from chloroplasts isolated by Conventional differential centrifugation. Barley chloroplast DNA size was sensibly lower (130 kpb) than lettuce chloroplast DNA (150 kpb). Chloroplast DNAs from the three barley cultivars showed similar restriction fragment patterns after digestion with: BamHI, EcoRI or HindIII. The lettuce chloroplast DNA restriction pattern was very different from the barley chloroplast DNA restriction pattern.     

Identification of Carnobacterium Species by Restriction Fragment Length Polymorphism of the 16S-23S rRNA Gene Intergenic Spacer Region and Species-Specific PCR

The genus Carnobacterium is currently divided into the following eight species: Carnobacterium piscicola, C. divergens, C. gallinarum, C. mobile, C. funditum, C. alterfunditum, C. inhibens, and C. viridans. An identification tool for the rapid differentiation of these eight Carnobacterium species was developed, based on the 16S-23S ribosomal DNA (rDNA) intergenic spacer region (ISR). PCR-restriction fragment length polymorphism (PCR-RFLP) analysis of this 16S-23S rDNA ISR was performed in order to obtain restriction profiles for all of the species. Three PCR amplicons, which were designated small ISR (S-ISR), medium ISR (M-ISR), and large ISR (L-ISR), were obtained for all Carnobacterium species. The L-ISR sequence revealed the presence of two tRNA genes, tRNA^Ala and tRNA^Ile, which were separated by a spacer region that varied from 24 to 38 bp long. This region was variable among the species, allowing the design of species-specific primers. These primers were tested and proved to be species specific. The identification method based on the 16S-23S rDNA ISR, using PCR-RFLP and specific primers, is very suitable for the rapid low-cost identification and discrimination of all of the Carnobacterium species from other phylogenetically related lactic acid bacteria.     

Sequence Characterization of a Unique Intergenic Spacer in Gadiformes Mitochondrial DNA

The nucleotide sequences of intergenic spacers located between the tRNA^Thr and tRNA^Pro genes in mitochondrial DNA of cod fishes (order Godiformes) were determined. Spacers from eight species representing two families of cod fishes were analyzed and found to vary in size from 25 to 99 bp. Each spacer sequence contains one or two copies of a conserved 17-bp motif. Four to five central nucleotides of this motif constitute a substitutional hot spot as observed from interspecific and intraspecific comparisons. The substitution rate of the spacer is approximately twice that of the variable part I of the mitochondrial DNA control region, making this sequence region interesting as a molecular marker in population studies or stock assessments of cod fishes. We propose that the spacer originated in a duplication event and evolved into a functional domain, perhaps by binding regulatory proteins. Accepted February 26, 1999     

中国桫椤科植物叶绿体trnL内含子和trnL-trnF基因间隔区序列的系统发育分析

运用对PCR产物直接测序和克隆后测序的方法测定了蚌壳蕨科1种和桫椤科11种(其中桫椤分别测定19株：小羽桫椤测定2株)植物的叶绿体trnL基因内含子和trnL-trnF基因间隔区序列。12种植物相应的长度介亍l004-l082之间,A T平均含量60．9％,G C平均含量39．1％。计算了不同种间以及种内不同个体间序列的碱基差别(转换值／颠换值)和Kimura遗传距离。序列数据经排列后分别进行最简约法、最大似然法和邻接法分析,结果显示：(1)白桫椤、海南白桫椤和大羽桫椤构成的分支最早和该科内其余植物组成的另一分支分歧,而后者又进一步分为刃个亚分支,分别和桫椤亚属、黑桫椤亚属对应,支持夏群的分类处理：(2)大桫椤～狭羽桫椤～毛轴桫椤～篦齿桫椤、多羽桫椤一白桫椤～海南白桫椤以及小羽桫椤一桫椤各自构成独立、自然的末端分支,再参照分支内植物间的遗传距蔼取值,建议将此3个末端分支依次归并为3种：大桫椤、白桫椤和桫椤;(3)白桫椤属在科内处于基部位置,桫椤属奇桫椤亚属和黑桫椤亚属为衍生分支,赞同Tryon关于桫椤科进化和囊群盖起源的假说。     

大麦花药培养中海藻酸钠作用的初步探讨

本试验以普通大麦（Hordeum vulgare L.）品种单核晚期的花药为材料,研究了他们在液体培养过程中海藻酸钠的作用。结果表明：在供试的4个海藻酸钠浓度（1、2、3、6%）中,以浓度为2-3%的效果最佳。当他们代替20-30%的Ficoll时,无论是用BAC3培养基,还是用MS培养基,培养基中加NAA,还是家2,4-D,都能诱导出愈伤组织,但以每升培养基加2mg NAA和1 mg 6-BA的效果最佳,在含这种激素的培养基上,一些基因型的出愈率相当高,供试基因型都在100%以上,而且有些花粉粒能直接发育成能正常发芽出苗的胚状体。Barley anthers at the late uninucleate stage were cultured in order to study the use of sodium alginate in liquidmedium.The results showed that, the effect of 2-3% (W/V) sodium alginate was best in 4 concentration studied (1%,2%,3%and 6%). When the 2-3% sodium alginate was substituted for 20-30% ficoll, the calli could be induced from anthers of all genotypes whether on BAC3 medium or MS medium and whether adding NAA or 2,4-Din medium.2mg/L NAA+1mg/L 6-BA added to medium could give better results and on the medium with these hormones the callus rate of all genotypes were above 100%. Some pollens could grow into embryoids directly.     

Identification of Heterotrophic Nanoflagellates by Restriction Fragment Length Polymorphism Analysis of Small Subunit Ribosomal DNA

Thirty clones derived from twenty isolates of heterotrophic nanoflagellates originating from a variety of marine and freshwater environments were examined by restriction fragment length polymorphism analysis of small subunit ribosomal RNA genes amplified by the polymerase chain reaction (riboprinting). The data were compared with light and electron microscopical identification of the isolates. On morphological criteria, sixteen of the thirty clones belonged to the genus Paraphysomonas De Saedeleer, seven to the genus Spumella Cienkowski, four to the genus Pteridomonas Penard and three to the genus Cafeteria Fenchel and Patterson. Among these taxa, eleven ribotypes were detected by analysis with the restriction enzymes Hinf I, Hae III, Sau3A I, and Msp I. Differentiation of nanoflagellate taxa by the riboprinting method supported taxonomic classification based on morphology at the generic and species level. The utility of the method for discriminating the 'naked' flagellates and for confirming the identity of polymorphic forms among species of Paraphysomonas is demonstrated.     

大麦胚性细胞悬浮系的建立及其生长特性

由春大麦品种“如车”种胚诱导的松脆型胚性愈伤组织经2个月的悬浮培养,成功建立分散性好、生长速度快的胚性细胞悬浮系。该系细胞直径为1-3mm,由富含淀粉粒的胚性薄壁细胞构成。经不同浓度2,4-D实验,发现2mg/L最适合该细胞系的生长。文中对成功建立大麦胚性细胞悬浮系的关键问题进行了讨论。     

人同源异型盒基因Hu-1位点的RFLPs研究

用12种限制性核酸内切酶分别对4至104条人类染色体进行酶谱分析,以寻找Hu-1基因附近的限制性酶切多态位点,仅发现了Bg 1Ⅱ酶的一个多态性位点。表现为Bg 1Ⅱ酶谱中除3.6kb片段外,杂合子型尚具6.3及6.6kb两种片段,而纯合子型则只有6.3或6.6kb一种片段。在所检查的104条染色体中6.6kb片段出现的频率为3.88％,而6.3kb片段出现的频率为96.12％。经统计分析,计算出Hu-1基因及其附近的核苷酸变异率为0.0017,大体上与人β珠蛋白基因簇及人α-1-抗胰蛋白酶基因的核苷酸变异率相似。文中对本工作的意义进行了初步讨论。