黑曲霉木质纤维素降解酶基因的可变剪接分析及验证

黑曲霉Aspergillus niger因能够产生大量的木质纤维素降解酶而在木质纤维素资源利用中发挥重要作用。目前,有关黑曲霉基因组中与木质纤维素降解相关的基因是否存在可变剪接的情况尚不清楚。本研究以黑曲霉CBS513.88菌株为研究对象,采用rMATS和ABLas两种方法对黑曲霉在葡萄糖为唯一碳源(G组)和小麦秸秆为唯一碳源(WS组)下的56个木质纤维素降解酶基因的可变剪接事件进行分析,并通过RT-PCR扩增和内含子特异性扩增对3个典型基因的可变剪接体进行了验证。结果表明,ABLas可变剪接分析算法相较于rMATS分析算法更为准确,ABLas分析算法显示G组和WS组共有21个木质纤维素降解酶基因出现了可变剪接,可变剪接类型以内含子保留(IR)为主,占所有可变剪接事件的82.85%。另外,G组和WS组发生可变剪接的木质纤维素降解酶基因也有所不同：G组发生可变剪接的基因为13个,WS组发生可变剪接的基因为14个,两组都发生可变剪接的基因为6个,这表明黑曲霉木质纤维素降解酶基因的可变剪接在不同生长条件下存在差异,另一方面,黑曲霉中众多可变剪接体的存在也为开发新型的木质纤维素降解酶资源提供基础。     

Phylogenetic relationships of Eurasian pines (Pinus, Pinaceae) based on chloroplast rbcL, MATK, RPL20-RPS18 spacer, and TRNV intron sequences

The sequence divergence of chloroplast rbcL, matK, trnV intron, and rpl20-rps18 spacer regions was analyzed among 32 Pinus species and representatives of six other genera in Pinaceae. The total aligned sequence length is 3570 bp. Of the four sequences examined, matK evolved much faster than rbcL in Pinus and in other Pinaceae genera. The two noncoding regions did not show more divergence than the two coding regions, especially within each Pinus subgenus. Phylogenetic analyses based on these four sequences gave consistent results and strongly supported the monophyly hypothesis for the genus Pinus and its two recognized subgenera. Pinus krempfii, the two-flat-needle pine endemic to Vietnam, was placed in subgen. Strobus and showed closer affinity to subsect. Gerardianae. The ancient character of sect. Parrya is further confirmed. However, monophyly of the sect. Parrya is not supported by our data. Among the Eurasian pines of subgen. Pinus, Mediterranean pines formed one clade and the Asian members of subsect. Sylvestres formed another. The Himalayan P. roxburghii showed considerable divergence from all the other hard pines from both regions. Pinus merkusii was distinctly separated from all the Asian members of subsect. Sylvestres. The implications of our results for Pinus classification are discussed.     

Molecular phylogenetics ofCochlearia (Brassicaceae) and allied genera based on nuclear ribosomal ITS DNA sequence analysis contradict traditional concepts of their evolutionary relationship

The systematics and phylogeny of the genusCochlearia and allied genera are unsettled. There are no clearly defined genera and subtribal structures to determine subtribeCochleariinae in respect to subtribeThlaspidinae. The use of morphological data, such as fruit form or embryo characters have resulted in contradictory taxonomic concepts in the past due to their homoplastic nature. We investigated all sections of genusCochlearia recognised in the most common concepts, as well as some genera such asIonopsidium, Bivonaea, Pastorea andThlaspi s. l. pro parte. Previous studies based on molecular data and morphological studies have shown close relationships between taxa from subtribeCochleariinae andThlaspidinae. The Internal Transcribed Spacer regions of the nuclear encoded ribosomal DNA operon and the plastidictrnL intron were sequenced from a number of genera. A molecular phylogeny was derived and compared to traditional classification systems. These data grouped sections ofCochlearia outside theCochlearial Ionopsidium core group and integrated them either closely to genusNoccaea in subtribeThlaspidinae (sect.Pseudosempervivum) or positioned them outside both theCochlearia core group and theThlaspi s. l. clade (sect.Hilliella). The molecular data indicate that subtribal arrangements in tribeLepidieae are artificial and do not reflect evolutionary history. The genusCochlearia is represented by sectionsCochlearia andGlaucocochlearia and the genusIonopsidium should be integrated intoCochlearia.     

Chloroplast DNA-relationship in palaeoaustralLopidium concinnum (Hypopterygiaceae, Musci). An example of stenoevolution in mosses Studies in austral temperate rain forest bryophytes 2

Evolutionary relationship between disjunct populations of the palaeoaustral moss taxonLopidium concinnum (Hypopterygiaceae) from New Zealand and southern South America were studied using non-coding chloroplast DNA sequences. No or only slight changes could be observed within the sequences oftrnT_UGU —trnL_UAA 5exon intergenic spacer,trnL_UAA intron andtrnL_UAA 3exon —trnF_GAA intergenic spacer. This indicates nearly no genetic divergence between extant New Zealand and Chilean populations, i.e. no significant differing pathways of evolution within the 80–60 million years of disrupted areas with interrupted gene flow. Molecular data support the idea of an old Gondwanan relict species of stenoevolutionary character. Ecological data on short-range dispersal strengthen this assessment.     

The evolutionary gain of spliceosomal introns: sequence and phase preferences

Theories regarding the evolution of spliceosomal introns differ in the extent to which the distribution of introns reflects either a formative role in the evolution of protein-coding genes or the adventitious gain of genetic elements. Here, systematic methods are used to assess the causes of the present-day distribution of introns in 10 families of eukaryotic protein-coding genes comprising 1,868 introns in 488 distinct alignment positions. The history of intron evolution inferred using a probabilistic model that allows ancestral inheritance of introns, gain of introns, and loss of introns reveals that the vast majority of introns in these eukaryotic gene families were not inherited from the most recent common ancestral genes, but were gained subsequently. Furthermore, among inferred events of intron gain that meet strict criteria of reliability, the distribution of sites of gain with respect to reading-frame phase shows a 5:3:2 ratio of phases 0, 1 and 2, respectively, and exhibits a nucleotide preference for MAG GT (positions -3 to +2 relative to the site of gain). The nucleotide preferences of intron gain may prove to be the ultimate cause for the phase bias. The phase bias of intron gain is sufficient to account quantitatively for the well-known 5:3:2 bias in phase frequencies among extant introns, a conclusion that holds even when taxonomic heterogeneity in phase patterns is considered. Thus, intron gain accounts for the vast majority of extant introns and for the bias toward phase 0 introns that previously was interpreted as evidence for ancient formative introns.     

Recent intron gain in elongation factor-1alpha of colletid bees (Hymenoptera: Colletidae)

We discovered the presence of a unique spliceosomal intron in the F1 copy of elongation factor-1alpha (EF-1alpha) restricted to the bee family Colletidae (Hymenoptera: Apoidae). The intron ranges in size from 101 to 1044 bp and shows no positional sliding. Our data also demonstrate the complete absence of this intron from exemplars representing all other bee families, as well as from close hymenopteran relatives. A review of the literature finds that this intron is likewise absent from all other arthropods for which data are available. This provides unambiguous evidence for a relatively recent intron insertion event in the colletid common ancestor and, at least in this specific instance, lends support to the introns-late hypothesis. The comparative distribution of this novel intron also supports the monophyly of Colletidae and the exclusion of the Stenotritidae from this family, providing an example of the potential of some introns to act as robust markers of shared descent.     

Hybridization and Rorippa austriaca (Brassicaceae) invasion in Germany

Introgressive hybridization between the invasive Rorippa austriaca and the native R. sylvestris in Germany has been studied using chloroplast DNA (trnL intron) and amplified fragment length polymorphism. Three hybrid zones between the invasive and native species were located in the Ruhr Valley (Mülheim) and at the River Main near Würzburg (Randersacker, Winterhausen). In each hybrid zone hybridization was indicated by additivity of region-specific amplified fragment length polymorphism markers proving independent hybridization events. The hybrids were either morphologically intermediate (R. x armoracioides) or were close to R. sylvestris. The trnL intron of R. austriaca is characterized by a species-specific deletion. This diagnostic chloroplast marker of R. austriaca was detected in three individuals of R. sylvestris providing evidence for introgression of the invasive chloroplast into the native species. Bidirectional introgression of R. austriaca markers into R. sylvestris and of R. sylvestris markers into R. austriaca was detected in the amplified fragment length polymorphism analysis. Some of the invasive R. austriaca populations showed high within-population variation. A possible association among introgression, within-population variation and invasion success is discussed. The morphologically intermediate hybrid R. x armoracioides is currently spreading in northern Germany. It forms large populations without its parent species R. austriaca and R. sylvestris. It is concluded that hybridization between invasive R. austriaca and native R. sylvestris may lead to the evolution of a new invasive species R. x armoracioides.