黄鳝二价染色体上地高辛配基标记随机引物原位DNA合成的研究

在黄鳝二价染色体上, 运用地高辛配基标记的随机引物原位DNA合成技术诱导出了明暗相间、基本稳定且类似于R带的带状结构。本文对该结果进行了较详细的分析和讨论。 Abstract:The dark bands alternating with light bands,relatively stable and R-band-like structure was showed on the pachytene bivalents of Rics-field eel(Monopterus albus Zuiew)by using the random-primed in situ digoxigenin-labelled DNA synthesis technique,and the results were analysed and discussed in detail.     

Evaluation of Genetic and Epigenetic Modification in Rapeseed （Brassica napus） Induced by Salt Stress

Salinity is an important limiting environmental factor for rapeseed production worldwide. In this study, we assessed the extent and pattern of DNA damages caused by salt stress in rapeseed plants. Amplified fragment length polymorphism （AFLP） analysis revealed dose-related increases in sequence alterations in plantlets exposed to 10-1000 mmol/L sodium chloride. In addition, individual plantlets exposed to the same salt concentration showed different AFLP and selected region amplified polymorphism banding patterns. These observations suggested that DNA mutation in response to salt stress was random in the genome and the effect was dose-dependant. DNA methylation changes in response to salt stress were also evaluated by methylation sensitive amplified polymorphism （MSAP）. Three types of MSAP bands were recovered. Type Ⅰ bands were observed with both isoschizomers Hpa Ⅱ and Msp Ⅰ, while type Ⅱ and type Ⅲ bands were observed only with Hpa Ⅱ and Msp Ⅰ, respectively. Extensive changes in types of MSAP bands after NaCI treatments were observed, including appearance and disappearance of type Ⅰ, Ⅱ and Ⅲ bands, as well as exchanges between either type Ⅰand type Ⅱ or type Ⅰ and type Ⅲ bands. An increase of 0.2-17.6% cytosine methylated CCGG sites were detected in plantlets exposed to 10- 200 mmol/L salt compared to the control, and these changes included both de novo methylation and demethylation events. Nine methylation related fragments were also recovered and sequenced, and one sharing a high sequence homology with the ethylene responsive element binding factor was identified. These results demonstrated clear DNA genetic and epigenetic alterations in planUets as a response to salt stress, and these changes may suggest a mechanism for plants adaptation under salt stress.     

蓖麻蚕DNA导入家蚕引起遗传变异的研究-基因组DNA的RAPD检测

借助于精子介导,在家蚕受精的过程中将蓖麻蚕DNA转入家蚕卵内,从它们后代获得了新的变异品系。本文采用RAPD技术对这些品系基因组DNA进行了分析。结果表明,所用50种10mer随机引物中有49个检测出DNA的多态性,统计分析图谱中各类扩增带,其中变异品系与其相应受体的差异带占其总带数的26-37%,提示外源DNA导入受体后引起后代基因组的显著变异,并对这些变异的意义了讨论。 Abstract:With the aid of domesticated silkworm sperms,eri silkworm DNA was transferred into domesticated silkworm eggs during insemination,and variant strains were obtained from the progenies.Genomes of three new strains were analyzed using RAPD assay.Polymorphic fingerprints were obtained from 49 out of 50 primers.Different kinds of amplified bands in RAPD patterns were calculated and analyzed,the variant bands between variants and their recipients counted for 26~37% of the total bands of each variant.The results indicated that exogenous DNA introduced into recipients induced remarkable variation in progeny genomes.The significance of the variation was discussed.     

有和无甘油的聚丙烯酰胺胶在检测突变时的差别

有文献报道在非变性的聚丙烯酰胺中加入甘油可提高SSCP检测的灵敏度。我们的实验结果建议研究者在进行SSCP筛查未知突变时最好采用不加甘油的非变性的聚丙烯酰胺胶,这既省力省钱,又灵敏。在判读SSCP胶时,千万不要看到在双链带位置有一条比正常迁移率慢的带就判定为插入突变。此时要判定突变的性质,最好测序。 Abstract：It was reported that glycerol in the non-denatured SSCP polyacrylamide gel could increase the sensibility of detecting mutation. We detected the mutation of PKD 1 gene in the patients with autosomal dominant polycystic kidney disease.PCR com bined with SSCP(single-strained conformation polymorphism),the non-denatured 10% polyacrylamide gel without glycerol or 10% polyacrylamide gel with 5% glycerol and DNA sequencing method were used.Our results showed that four single strand b ands were found in the non-denatured polyacrylamide gel without glycerol while t wo single strand bands were found in the polyacrylamide gel with glycerol in the same patient.Sequence showed there is a deletion of G in one DNA molecular and a G→A substitution in another DNA molecular in the patient with abnormal shift SSCP bands.Therefore, our experiment suggested that non-denat ured polyacrylamide gel was better than the polyacrylamide gel with glycerol in detection mutation,and it will save labor and money.It also suggeste d that one basedeletion can cause a slow double-strand DNA following the normal double strand band,which was caused by the heterogeneous DNA molecule formed bet ween the normal DNA strand and the one base deletion DNA strand with the protrud ing base.Our results suggest that when judging mutation in SSCP gel,it is not re liable to decide that mutation is inversion according to slow mobility in the ge l,and when the characteristic of mutation need to be judged,it must be sequenced .     

Karyotyping of Brassica napus L. Based on C0t-1 DNA Banding by Fluorescence In Situ Hybridization

In order to precisely recognize and karyotype Brassica napus L. chromosomes, C0t-1 DNA was extracted from its genomic DNA, labeled with biotin-1 1-dUTP and in situ hybridized. The hybridized locations were detected by Cy3-conjugated streptavidin. Specific fluorescence in situ hybridization （FISH） signal bands were detected on all individual chromosome pairs. Each chromosome pair showed specific banding patterns. The B. napus karyotype has been constructed, for the first time, on the basis of both Cot-1 DNA FISH banding patterns and chromosome morphology.     

Inhibition of shoot induction by 5-azacytidine and 5-aza-2′-deoxycytidine in Petunia involves DNA hypomethylation

Shoot bud regeneration from Petunia leaf disks was inhibited when they were cultured with the demethylating agents, 5-azacytidine (AzaC) and 5-aza-2′-deoxycytidine (AzadC), in shoot induction (SI) medium. Explants induced shoot primordia if they were transferred after 1 week from the medium containing the drugs to medium without drugs. The fresh weight of leaf disks cultured on SI medium for 2 weeks in the presence of the drugs was 60–80% lower when compared to control shoot-forming cultures. Internode length was reduced when shoots were transferred to phytohormone-free Murashige and Skoog medium containing the drugs. However, no other morphological abnormalities were seen in these shoots, even at 20 μm AzaC or 5 μm AzadC. Coupled restriction enzyme digestion (with HpaII and MspI) and random amplification of genomic DNA was performed to detect the level of methylation of CCGG sites in the DNA of the explants exposed to AzaC and AzadC. Over 15 amplified bands were detectable in the control. Five of these bands were absent in the amplified products when digested DNA from the drug-treated explants was used as the template, showing that hypomethylation of DNA had occurred. This suggests that inhibition of shoot bud formation in the presence of the drugs AzaC and AzadC may be due to the altered methylation status. Received: 7 January 1997 / Revision received: 17 February 1997 / Accepted: 1 March 1997     

Identification and isolation of Mu-flanking fragments from maize

Transposable elements have been utilized as mutagens to create mutant libraries for functional genomics.Isolation of genomic seg-ments flanking the insertion Mutator (Mu) is a key step in insertion mutagenesis studies.Herein,we adopted a modified AFLP method to identify and isolate Mu-flanking fragments from maize.The method consists of the following steps: 1) double-digestion of genomic DNA with Bgl ⅡMsp Ⅰ and ligation of digested fragments to the Bgl Ⅱ- and Msp Ⅰ-adaptors; 2) enrichment of a subset of Bgl Ⅱ/Msp Ⅰ fragments followed by selective amplification of the Mu-flanking fragments; 3) simultaneous display of AFLP bands derived from the flanking re-gions for both insert and native Mu transposons; 4) identification and isolation of AFLP bands resulting from Mu insertions by comparing the banding profiles between Mu-induced mutants and their parental lines; and 5) confirmation of flanking fragments related to these Mu insertions.Using this approach,we have isolated flanking fragment(s) resulting from Mu insertion for every Mu-indueed mutant,and one such fragment,M196-FF,is found to contain a partial sequence of the DNA topoisomerase Ⅰ gene Topl.Moreover,the modified AFLP method including all restriction enzymes,adaptors and primers has been optimized in this study.The modified AFLP method has been proved to be simple and efficient in the isolation of Mu-flanking fragments and will find its usefulness in the functional genomics of maize.     

Bachelor groups in primate multilevel society facilitate gene flow across fragmented habitats

In the face of ongoing habitat fragmentation,many primate species have experienced reduced gene flow resulting in a reduction of genetic diversity,population bottlenecks,and inbreeding depression,including golden snub-nosed monkeys Rhinopithecus roxellana.Golden snub-nosed monkeys live in a multilevel society composed of several 1 male harem units that aggregate to form a cohesive breeding band,which is followed by one or more bachelor groups composed of juvenile,subadult,and adult male members.In this research,we examine the continuous landscape resistance surface,the genetic diversity and patterns of gene flow among 4 isolated breeding bands and 1 all-male band in the Qinling Mountains,China.Landscape surface modeling suggested that human activities and ecological factors severely limit the movement of individuals among breeding bands.Although these conditions are expected to result in reduced gene flow,reduced genetic diversity,and an increased opportunity for a genetic bottleneck,based on population genetic analyses of 13 microsatellite loci from 188 individuals inhabiting 4 isolated breeding bands and 1 all-male band,we found high levels of genetic diversity but low levels of genetic divergence,as well as high rates of gene flow between males residing in the all-male band and each of the 4 breeding bands.Our results indicate that the movement of bachelor males across the landscape,along with their association with several different breeding bands,appears to provide a mechanism for promoting gene flows and maintaining genetic diversity that may counteract the otherwise isolating effects of habitat fragmentation.     

Loss of Genetic Diversity of Domesticated Panax notoginseng F H Chen as Evidenced by ITS Sequence and AFLP Polymorphism： A Comparative Study with P. stipuleanatus H T Tsai et K M Feng

In the present study, we evaluated the genetic diversity of Panax notoginseng F H Chen, a domesticated species, and P. stipuleanatus H T Tsai et K M Feng, an endangered wild species in southeastern Yunnan and adjacent areas in Vietnam, using sequences of the internal transcribed spacer (ITS) regions of nuclear ribosomal DNA and amplified fragment length polymorphism (AFLP) markers. Twenty-four accessions from three plantations of P. notoginseng and 51 samples from eight populations of P. stipuleanatus were assayed. A total of 694 bp of partial sequences of 18S, ITS 1, 5.8S, ITS2, and partial sequences of 26S were obtained. No sequence variation was detected within P. notoginseng and nine sites (1.30%) were variable in P. stipuleanatus. Two-thirds of the variable sites were found between Langqiao and other populations. In P. notoginseng, four pairs of AFLP primer combinations generated 312 bands, of which 240 (76.9%) were polymorphic and 60.15% of the polymorphisms were harbored within plantations. Approximately 41.0% and 66.9% of bands were polymorphic in population D7 and 5589, respectively. In P.stipuleanatus, the same four primer combinations produced 346 bands, of which 334 (96.5%) were polymorphic and approximately 62.14% of polymorphisms were maintained within populations. Considerable variations were observed. The percentage of polymorphic bands ranged from 50.2% to 84.9% and the average over populations was 70.9%. Cluster analysis did not show correlation of genetic differentiation with the distinctive leaf morphology of P. stipuleanatus (i.e. one form with bipinnatifid leaflets and the other with undivided leaflets). Because over 40% of genetic variations were maintained among populations and because of the very restricted distribution of P. stipuleanatus, all natural populations of this species should be conserved in situ. Considering that there are variations in P. notoginseng within and among plantations, we suggest establishing a genetic resource conservation garden or reintroducing P. notoginseng into its native habitats in southwestern China. Such reintroduction should be carefully executed after large-scale screening of genetic variation within the species.     

小麦杂交种与亲本发育早期种子的基因表达差异及其与杂种优势关系的初步研究

为深入探讨小麦杂种优势形成的分子机理,选取3个冬小麦品种(系)为一组亲本,3个为另一组亲本,配制了正反交18个杂交组合,以授粉后6d的杂交和自交种子为材料,应用mRNA差异显示技术(DDRT—PCR)研究了小麦杂交当代种子与其亲本自交种子基因的表达差异,并与杂种优势进行相关分析。为降低DDRT—PCR技术假阳性的不利影响,对每个引物组合均作了两次PCR扩增,在处理数据时,仅统计能重复出现的条带。结果发现：杂交种和亲本之间的基因表达模式有8类共15种：(1)单亲沉默型(2种),(2)单亲一致型(2种),(3)正交或反交沉默型(2种),(4)正交或反交特异型(2种),(5)正交或反交单亲一致型(4种),(6)杂交种特异型(1种),(7)双亲共沉默型(1种),(8)表达一致型(1种)。分析发现,小麦杂交种和亲本间存在显著的表达差异。在差异表达类型中,杂交种特异型和双亲共沉默型比例最低。对上述15种表达模式与杂种优势进行相关分析,结果表明,表达一致型与各产量性状杂种优势之间的相关均不显著,说明杂种优势是由某些有表达差异的基因造成。9个产量性状均能检测到一种以上与其显著或极显著相关的基因表达模式,有些性状受正负相关效应的共同影响;沉默型(包括单亲沉默型、正交或反交沉默型和双亲共沉默型)和正交或反交单亲一致型在杂种优势形成中发挥重要作用。这些研究表明,在种子发育早期,基因的差异表达与杂种优势形成之间可能存在较为复杂的关系。     

Oligoclonal IgG bands with and without measles antibody activity in sera of patients with subacute sclerosing panencephalitis (SSPE)

Oligoclonal IgG bands from SSPE sera were isolated by combination of Protein A-Sepharose 4B column and preparative isoelectric focusing gel procedures. Each eluted fraction, when examined in analytic IEF, showed two or three individual bands with isoelectric points close to one another, compared to approximately fifteen IgG bands seen in whole serum. When the bands were tested for measles antibody activity in immunofixation with measles virus followed by peroxidase staining, the bands eluted in pH region 8.5 to 9.3 were found to be measles specific, whereas those in pH 7.0 to 8.4 lacked significant measles activity. When eluted fractions containing groups of bands were absorbed with measles virus, the bands in pH region 8.5 to 9.3 were removed, whereas those in pH 7.0 to 8.4 region remained unchanged; this indicated that a number of oligoclonal IgG bands without measles virus activities are present in SSPE. The bands lacking measles-specific activity may be synthesized against other infectious agents or they may represent nonspecific activation of B cell clones.     

Hepatitis B surface antigen polypeptides: artifactual bands in sodium dodecyl sulfate-polyacrylamide gel electrophoresis caused by aggregation.

Hepatitis B surface antigen, subtype ad, was purified and studied by sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing conditions. Two major bands with molecular weights of 23,500 and 27,500 and several weaker bands with higher molecular weights were observed. When the low-molecular-weight bands and the group of high-molecular-weight bands were excised from the gel, eluted, and reelectrophoresed, neither the low-molecular-weight bands nor the high-molecular-weight bands ever appeared alone, but both high- and low-molecular-weight bands always appeared. It was concluded that the apparently high-molecular-weight bands represented aggregates of the two small polypeptides whose monomers formed the major bands. The preparation thus contained only two polypeptides.     

玉米杂种与亲本穗分化期功能叶基因差异表达与杂种优势

为探讨玉米杂种优势的分子机理,以10个玉米自交系及其组配的38个杂交种为材料,利用cDNA-AFLP技术,分析杂种与亲本在玉米雌穗小穗分化期功能叶片的基因差异表达类型与主要农艺性状的杂种表现及杂种优势的关系。研究表明：（1）杂种的基因相对于其双亲,存在质和量的表达差异,其中质的差异表达类型包括：单亲沉默表达,双亲沉默表达,亲本显性表达和杂种特异表达等类型。（2）在雌穗小穗分化期,同一差异表达类型中不同杂交组合间差异很大;从总体平均看,杂种特异表达类型占25.22%,亲本显性表达类型占21.46%,双亲沉默表达类型占8.27%,单亲沉默表达类型占33.49%。（3）单亲沉默表达与株高的杂种表现呈显著正相关;双亲沉默表达与穗粗的杂种优势呈显著负相关,显性表达与行粒数和单株粒重的杂种优势呈显著负相关,其余表达类型与所有农艺性状杂种表现及杂种优势均不相关,并对结果进行了讨论。     

Infrared spectroscopic studies on interactions of water and carbohydrates with a biological membrane

Infrared spectroscopy was used to investigate the changes in bands assigned to phospholipids and proteins in dehydrated and rehydrated sarcoplasmic reticulum. The changes in CH₂ and CH₃ stretching bands, amide bands, and phosphate stretching bands are similar to shifts in frequency seen for those bands in phospholipid and protein preparations during thermotropic phase transitions and hydration. IR studies on dry trehalose-sarcoplasmic reticulum mixtures show similar results; with increasing trehalose concentration in the dry mixtures, amide and phosphate bands shift to frequencies characteristic of hydrated samples. Changes in bands assigned to OH deformations in the trehalose suggest that the interaction between the carbohydrate and membrane is by means of hydrogen bonding between these OH groups and membrane components.     

Continued use of soft‐metal bands on gulls in North America reduces the value of recovery data

Use of soft‐metal (aluminum alloy) bands on gulls (Laridae) is known to result in high rates of band loss and, as a result, hard‐metal (monel, incoloy, or stainless steel) bands are superior for most studies. However, the U.S. Bird Banding Laboratory (BBL) and the Canadian Wildlife Service Bird Banding Office continue to issue soft bands for use on gulls, and the BBL does not make specific recommendations about use of hard bands so many banders continue to use soft bands. For wholly marine species of gulls banded in North America since 1996, ～20% have been banded with soft bands; the proportion of soft bands used on partially freshwater gulls was ～70% up to 2009, but has since fallen to 40%. Using hierarchical Bayesian models in program MARK, we analyzed recovery data for three gull species and found that estimates of annual survival rates derived from soft bands (0.68–0.81) were lower than those derived from hard bands (0.85–0.96). Comparison of survival rates of Herring Gulls (Larus argentatus) in the Great Lakes basin and on the Atlantic coast provided no evidence that soft bands last longer in freshwater than saltwater. Band loss compromises many types of studies, including those assessing the possible effects of climate change. We recommend that use of soft bands on gulls be discontinued, and that banders be required to use hard bands on these species in the future. The same consideration applies to other long‐lived species, including some waterfowl and all albatrosses, pelicans, cormorants, shearwaters, petrels, terns, shorebirds, and alcids. Use of hard bands should be based on expectations about a species’ longevity and evidence of band wear, rather than on whether or not it occurs in saltwater.     

正常与感染粘孢子虫鲫鱼主要组织可溶性蛋白质电泳比较

采用SDS-聚丙烯酰胺凝胶电泳技术,对正常和感染粘孢子虫的鲫鱼动脉球、肝脏、鳃、脑、肠、肌肉、眼、鳍8种组织可溶性蛋白质进行分析比较。结果表明：病鱼的动脉球、肝脏、鳃、脑、肠5种组织的蛋白质电泳谱带与正常鱼相比有较大变化,动脉球缺失1条谱带同时又诱导出1条新谱带;肝脏有9条谱带缺失;鳃有2条谱带缺失的同时诱导出3条新谱带;脑中新增3条谱带;肠中有5条谱带缺失同时产生3条新谱带。这些变化可作为辅助疾病诊断的生化指标,为从生化水平上揭示该病的致病机理及开展有效的早期防治研究奠定基础。     

Tissue-, age-, and stage-specific patterns of protein synthesis during the development of Drosophila melanogaster.

Patterns of polypeptide synthesis in the imaginal and larval tissues of wild-type larvae of different ages have been examined using a one-dimensional polyacrylamide gel electrophoresis and autofluorography. Tissue-, age, and stage-specific patterns of polypeptide bands have been observed and characterized. There are one or two bands unique to larval tissues and 13 bands unique to imaginal discs and brain. Each tissue has its own characteristic band pattern. The 160-hr wing disc contains all 13 disc and brain-specific bands plus another eight unique bands. The 160-hr leg disc contains 11 disc- and brain-specific bands plus another six unique bands. The 160-hr eye-antennal disc contains eight disc- and brain-specific bands. There are five bands common to all imaginal tissues at all times but not found in any of the larval tissues. There appears to be a much greater difference between the same tissue at different ages than there is between different tissues at the same age. There is also a marked change in the total body proteins extracted from different developmental stages. Eight (11%) of the bands detected appear to be common to all tissues at all times, while 13 bands (18%) appear to be specific to pairs of imaginal discs. In addition, there are major differences in the patterns observed between pulsed and pulse-chased animals. These results are discussed in relation to the concept of differential gene activity.     

Precursor Products Found in Formaldehyde-fixed Lysates of BHK-21 Cells Infected with Pseudorabies Virus

Lysates of BHK-21 cells, infected with pseudorabies and labeled with ³H-thymidine, were treated with formaldehyde and centrifuged in preformed CsCl gradients. Five peaks, designated bands A to E, were detected and shown to contain the following virus-specific products: band A, aggregates of virions; band B, virions; band C, empty capsids and nucleocapsids; bands D and E, small nucleoid-like particles. All five bands contained viral deoxyribonucleic acid and virous-specific antigens. Deoxyribonuclease treatment of the lysates before centrifugation resulted in the complete loss of ³H-thymidine-labeled material from bands D and E; bands B and C were unaffected. Pulse-chase studies showed that ³H-thymidine can be rapidly chased into the virus-specific products sedimenting in bands D and E. Further chase resulted in the gradual loss of label from bands D and E with a concomitant increase in the amount of label in bands A, B, and C. These results indicate that the virus-specific products in bands D and E are precursors of pseudorabies virions and may represent the nucleoid of the virion.     

Mourning dove reporting probabilities for web-address versus toll-free bands

Federal migratory bird bands have been inscribed with a postal address and toll-free telephone number (toll-free bands) since 1995 for their reporting, but in 2007 the postal mail address was replaced with an internet address (web-address bands). The reporting rate of web-address bands is unknown, and knowledge of band reporting probabilities is an integral part of bird band-recovery studies designed to obtain unbiased estimates of harvest rates and recovery rates if use of band types differs across space and time. We estimated web-address band reporting probabilities for mourning doves (Zenaida macroura) in the United States by comparing the relative recovery rate (ratio) of web-address to toll-free bands and applying this as an adjustment factor to the known reporting probabilities of toll-free bands. Cooperators banded 132,386 doves representing 57,982 (44%) web-address and 74,404 (56%) toll-free bands in July and August during 2008–2010 across 34 states, 12 regions, and 3 management units. Of the birds banded, 4,642 (3.5%) were direct recoveries, as a result of being shot, and all but 3 were reported via telephone or internet. Nationally, internet reporting accounted for 16.3% of toll-free bands and 42.9% of web-address bands recovered. Web-address bands had a greater recovery rate than toll-free bands (ratio = 1.081, SE = 0.027). We found some evidence of spatial variation in ratio estimates at management unit and regional scales, but the variation was primarily at the regional level where it was not great (range = 1.015–1.213) and statistical support for this variation was not particularly strong (SE = 0.129). Nationally, the web-address band reporting probability was 0.535 (SE = 0.021), an absolute increase of 0.040 compared to the reporting probability of toll-free bands. The additional reporting probability of web-address bands relative to toll-free bands must be accounted for in band-recovery studies used to obtain estimates of harvest rates and recovery rates if use of band types differs across space and time for mourning doves and possibly other species. © 2011 The Wildlife Society.