A rapid single-stranded cloning strategy for producing a sequential series of overlapping clones for use in DNA sequencing: application to sequencing the corn mitochondrial 18 S rDNA

A simple new procedure was described for producing a sequential series of overlapping clones for use in DNA sequencing. The technique used single-stranded M13 DNA and complementary DNA oligomers to form specific cleavage and ligation substrates. It was, therefore, independent of the sequence of the DNA cloned into the vector. Deletions of varying sizes were generated from one end of the insert through the 3' to 5' exonuclease activity of T4 DNA polymerase. The approximate size of the deletion and therefore the starting point for DNA sequencing could be estimated by electrophoresis of the subcloned phage DNA on a agarose gel. This greatly reduced the number of templates that must be sequenced to obtain a complete sequence. The entire procedure could be carried out in one tube in less than a day. The procedure was used to subclone and sequence the maize mitochondrial 18 S rDNA and 5' flanking region (2622 bases) in less than a week. Other applications of oligomers and single-stranded DNA in the construction of insertions, deletions, and cDNAs are discussed.     

Relationships betweenOryza species (Poaceae) based on 5S DNA sequences

Relationships between 9Oryza species, covering 6 different genomes, have been studied using hybridization and nucleotide sequence information from the5S Dna locus. Four to five units of the major size class of 5S DNA in each species, 55 units in all, were cloned and sequenced. Both hybridization and sequence data confirmed the basic differences between the A and B, C, D genome species suggested by morphological and cytological data. The 5S DNA units of the A genome species were very similar, as were the ones from the B, C, and D genome-containing species. The 5S DNA ofO. australiensis (E genome) grouped with the B, C, D cluster, while the units ofO. brachyantha (F genome) were quite different and grouped away from all other species. 5S DNA units fromO. minuta, O. latifolia, O. australiensis, andO. brachyantha hybridized strongly, and preferentially, to the genomic DNA from which the units were isolated and hence could be useful as species/genome specific probes. The 5S DNA units fromO. sativa, O. nivara, andO. rufipogon provided A genome-specific probes as they hybridized preferentially to A genome DNA. The units fromO. punctata andO. officinalis displayed weaker preferential hybridization toO. punctata DNA, possibly reflecting their shared genome (C genome).     

DNA sequence analysis of a 5.27-kb direct repeat occurring adjacent to the regions of S-episome homology in maize mitochondria.

The DNA sequence of the 5270-bp repeated DNA element from the mitochondrial genome of the fertile cytoplasm of maize has been determined. The repeat is a major site of recombination within the mitochondrial genome and sequences related to the R1(S1) and R2(S2) linear episomes reside immediately adjacent to the repeat. The terminal inverted repeats of the R1 and R2 homologous sequences form one of the two boundaries of the repeat. Frame-shift mutations have introduced 11 translation termination codons into the transcribed S2/R2 URFI gene. The repeated sequence, though recombinantly active, appears to serve no biological function.     

Kinetic evidence for involvement of two cytochrome b-563 hemes in photosynthetic electron transport

The effect of 2-(n-heptyl)-4-hydroxyquinoline N-oxide (HQNO) on the kinetics of cytochrome b-563 and cytochrome c² turnovers following single-turnover flashes was measured in isolated heterocysts. Low concentrations of HQNO (below 3 μM) blocked reoxidation of cytochrome b-563, whereas higher concentrations (above 5 μM) resulted in additional inhibition of cytochrome b-563 oxidation and also inhibited reduction of cytochrome b-563 and cytochrome c. Similar effects on cytochrome b-563 reduction and reoxidation were obtained with a combination of 5 μM HQNO and 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (1–7 μM). In HQNO-inhibited heterocysts, cytochrome c reduction following a flash occurred in three phases with half-times of 0.5, 2.8 and 45 ms. The second phase nearly equalled the cytochrome b-563 reduction in half-time and magnitude. In the presence of HQNO, the reoxidation of cytochrome b-563 following two closely spaced actinic flashes displayed biphasic kinetics. The two phases correspond to reoxidation of cytochrome b-563 in which one or both of the cytochrome b-563 hemes in the cytochrome b–f complex are reduced. These results are interpreted in terms of a Q-loop in which HQNO, at low concentrations, blocks the site of rapid cytochrome b-563 reoxidation and at higher concentrations, also inhibits the site of electron donation by plastoquinol to the cytochrome b-f complex.     

我国微茎类吸虫研究:ⅩⅢ.微茎类吸虫成虫结构特征排序

对我国52种微茎类吸虫的18项成虫形态学特征进行主成分分析,结果表明:卵巢位置、子宫延伸位置等7项性状对第一主成分贡献较大,提示描述器官位置的指标是重要的分类依据。52个虫种在前三个主成分上的排序图显示应将其划分成4个亚科。     

我国微茎类吸虫的研究Ⅻ.类茎属及多黄属二新种报道：复殖目：微茎科

本文报道在湛江市附近海域海鸟体内获得的两种吸虫,经鉴定为新种,命名为巨口类茎吸虫,新种Ｍｉｃｒｏｐｈａｌｌｏｉｄｅｓ ｍａｃｒｏｓｔｏｎｒｓ ｓｐ．ｎｏｖ．,珊瑚多黄吸虫,新种Ｍｕｌｔｉｖｉｔｅｌｌｕｓ ｃｏｒａｌｉｕｓ ｓｐ．ｎｏｖ．     

以化学纯饲料饲养北京的桃蚜

应用修改后的Dadd和Mitter(1966)全纯饲料配方配制成人工饲料饲养定居在北京温室烟草上的桃蚜 Myzus persicae可完成生活史井连续饲养3代。本文描述饲料配制、饲养和取食量测定的方法。这3代初羽化无翅孤雌胎生雌蚜的平均体重分别为:440±90.7μg,264±104.9μg和312±127.9μg。用放射性同位素稀释法测定取食量的结果得悉若虫期的总取食量每蚜约为1.74μg,相当于1.16μl。     

合肥秋冬季茶园天敌对假眼小绿叶蝉和茶蚜的空间跟随关系

为了合理保护和利用天敌及科学地选取抽样方法,开展了合肥地区秋冬季4个品种茶园假眼小绿叶蝉Empoasca vitisGothe和茶蚜Toxoptera aurantii Boyer与其天敌之间空间关系研究,运用地学统计学方法求得天敌和害虫各自的变程,用灰色关联度分析方法分析害虫与天敌变程的关联度,关联度值越大的天敌在空间上对害虫的跟随关系越密切。分析了2010年9月28日至11月25日期间假眼小绿叶蝉和茶蚜数量最少的舒茶早茶园和二种害虫数量最多的平阳特早茶园天敌对害虫空间上的跟随关系,结果表明,二种茶园的假眼小绿叶蝉和茶蚜及其4种主要天敌均为聚集分布,舒茶早茶园与假眼小绿叶蝉空间上跟随关系密切的前二位天敌是斜纹猫蛛Oxyopes sertatus L.Koch(0.8594)和草间小黑蛛Erigonidium graminicolum Sundevall(0.8397),与茶蚜空间上跟随关系密切的前二位天敌是草间小黑蛛(0.7448)和斜纹猫蛛(0.7433);平阳特早茶园与假眼小绿叶蝉空间上跟随关系密切的前二位天敌是八斑球腹蛛Theridion ocomaculatum Bose.et Str(0.8207)和斜纹猫蛛(0.8104),与茶蚜空间上跟随关系密切的前二位天敌是八斑球腹蛛(0.8324)和斜纹猫蛛(0.7730)。其中,11月25日4种茶园假眼小绿叶蝉和茶蚜数量均较多。分析了该日另外二个茶树品种福云六号和龙井长叶茶园假眼小绿叶蝉和茶蚜与其天敌的空间关系,结果表明,二种茶园假眼小绿叶蝉和茶蚜及其天敌均为聚集分布,福云六号茶园与假眼小绿叶蝉变程值(3.8182)最接近的天敌是斜纹猫蛛(4.7222),与茶蚜变程值(6.5854)最接近的天敌是斜纹猫蛛(4.7222);龙井长叶茶园与假眼小绿叶蝉变程值(1.0000)最接近的天敌是八斑球腹蛛(1.0000),与茶蚜变程值(4.5000)最接近的天敌是斜纹猫蛛(7.6316)。总之,秋冬季4个品种茶园斜纹猫蛛在空间上是假眼小绿叶蝉和茶蚜跟随关系最密切的天敌,其次是八斑球腹蛛和草间小黑蛛。     

李肖叶甲成虫数量及三维空间格局动态

对合肥地区李肖叶甲成虫数量动态及三维空间格局进行研究,结果表明该成虫5月下旬初见,6月4日至7月16日为发生高峰期,9月上旬仍有零星虫口。6月4日至9月10日种群增长模型为y=1692.2258e-0.0545t,6月4日至8月27日树冠上部部位增长模型为y=205.60e-0.0523t,树冠西部部位的增长模型为y=257.13e-0.0505t。样地中的李肖叶甲5月21日至8月13日半变异函数方程是y=0.0709x3-10.479x2+391.67x-300.71、y=-0.0122x3+1.1201x2-19.781x+317.84、y=-0.0013x3+0.1613x2-4.4862x+67.363、y=-0.0016x3+0.9177x2-11.495x+551.94、y=-0.0029x3+0.3034x2-7.5906x+103.37和y=-0.0002x3+0.0172x2-0.4975x+13.691,变程在20.3938—65.0289之间,均为聚集格局,聚集强度指标表明也均为聚集格局;树冠东、西、南、北方位的水平分布5月21日至8月13日的C值均大于1,I值均大于0,CA值均大于0,Iw值均大于1,为聚集格局;树冠上、中、下方位的垂直分布6月4日至8月13日的C值均大于1,I值均大于0,CA值均大于0,Iw值均大于1,为聚集格局;5月21日至8月13日的树冠东、南、西、北、上、中、下部位的C值均大于1,I值均大于0,CA值均大于0,Iw值均大于1,三维分布均为聚集格局。用Iwao公式计算的结果与聚集强度指标分析的结果一致。6月4日—8月13日林间李肖叶甲的种群聚集均数λ值均大于2,聚集是李肖叶甲本身的行为所致。     

城市扩张与耕地保护耦合对陆地生态系统碳储量的影响——以湖北省为例

城市扩张对生态系统的影响可以分为城市扩张对生态用地的直接挤占以及为补充城市扩张占用的耕地对生态用地二次挤占两个连锁的过程。以往的研究大多只关注城市扩张对生态系统的直接影响,忽略了城市扩张与耕地保护耦合的间接影响。针对这一问题,采用GIS空间分析方法和InVEST模型定量评估了2000年至2015年湖北省城市扩张与耕地保护耦合对陆地生态系统碳储量的影响。结果显示:(1)城市扩张与耕地保护耦合导致湖北省陆地生态系统碳储量减少40.90 Tg;(2)由城市扩张与耕地保护耦合导致的陆地生态系统碳储量的减少量中,耕地保护传导作用间接导致的占比31%。结果表明,城市扩张与耕地保护的耦合是导致生态系统碳储量减少的主要原因之一。忽视城市扩张通过耕地保护的传导作用对生态系统碳储量的间接影响,会导致城市扩张对陆地生态系统碳储量的影响被严重低估。