Isolation and Characterization of Transcriptionally Active Ty1-copia Retrotransposons in Fragaria × ananassa

One possible mechanism suggested for somaclonal variation is the activation of transposable elements. The activation of retrotransposons by stresses and external changes is commonly observed in plants. In previous study, we isolated the reverse transcriptase （RT） gene sequences of Ty 1-copia retrotransposons from tissue culture strawberry （Fragaria x ananassa） plant, but not the transcriptionally active sequence. For further understanding the relationship between retrotransposon and somaclonal varation, in this study, we isolated the transcriptionally active RT gene sequences from strawberry plants subjected to different abiotic stresses. These retrotransposons were activated by spraying strawberry leaves with 2 mmol L^-1 salicylic acid （SA）, 50 mmol L^-1 methyl jasmonate （MeJA）, 50 mmol L^-1 abscisic acid （ABA）, 50 mmol L^-1 2,4- dichlorophenoxyacetic acid （2,4-D） or by inducing callus growth in 2 types of MS media： first medium supplemented with 0.5 mg L^-1 6-benzylaminopurine （6-BA）, 0.5 mg L^-1 gibberellic acid （GA3）, 1.0 mg L^-1 thidiazuron （TDZ）, and 0.1 mg L^-1 2,4-D, and the second medium supplemented with 0.5 mg L^-1 6-BA, 0.5 mg L^-1 GA3, 2.0 mg L^-1 TDZ, and 0.02 mg L^-1 indole butyric acid （1BA）. Analysis of gene sequences of 17 RTs revealed that none of them contained stop codons and/or indels disrupting the reading frame. These different stress-origin transcriptionally active RTs were remarkably similar to each other- FATEXP2-8 and FATEYS9-7 showed 100% sequence identity. Analysis of pylogenetic of these transcriptionally active RTs and the RT sequences from genome showed that there were close phylogenetic relationships of most of the transcriptionally active RTs. The results of this study have contributed to the background information necessary for future studies for evaluating the relationship between retrotransposons and somaclonal variation.

Impact of Low-Energy Ion Beam Implantation on the Expression of Ty1-copia-like Retrotransposons in Wheat(Triticum aestivum)

Retrotransposon-like elements are major constituents of most eukaryotic genomes. For example, they account for roughly 90% of the wheat （Triticum aestivum） genome, Previous study on a wheat strain treated by low-energy N^＋ ions indicated the variations in AFLP （Amplified Fragment Length Polymorphism ） markers, One such variation was caused by the re-activation of Tyl-copia-like retrotransposons, implying that the mutagenic effects of lowenergy ions might work through elevated activation of retrotransposons, In this paper an expression profile of Tyl-copia-like retrotransposons in wheat treated by low-energy N^＋ ions is reported, The reverse transcriptase （RT） domains of these retrotransposons were amplified by reverse-transcriptional polymerase chain reaction （RT-PCR） and sequentially cloned, 42 and 65 clones were obtained from the treated （CL） and control materials （CK）, respectively, Sequence analysis of each clone was performed by software. Phylogeny and classification were calculated responding to the sequences of the RT domains. All the results show that there is much difference in the RT domain between the control sample and the treated sample, Especially, the RT domains from the treated group encode significantly more functional ORF （open reading frames） than those from the control sample, This observation suggests that the treated sample has higher activation of retrotransposons, possibly as a consequence of low-energy ion beam irradiation, It also suggests that retrotransposons in the two groups impact the host gene expression in two different ways and carry out different functions in wheat cells.

Isolation and Characterization of Copia-like Retrotransposons from 12 Sweet Orange （Citrus sinensis Osbeck） Cultivars

As the largest transposable element in the plant genome, retrotransposons are thought to be involved in citrus genetic instability and genome evolution, especially in sweet orange, which is prone to bud mutation. In the present study, the presence of copia-like retrotransposons, their heterogeneity, genomic distribution, and transcriptional activities in Citrus were investigated in 12 sweet orange （Citrus sinensis Osbeck） cultivars using a PCR assay designed to detect copia-like reverse transcriptase （RT） sequences. Twelve amplification products from each cultivar were cloned and sequenced. The cloned sequences showed great heterogeneity, except “Dream” navel and “Hamlin”, both of which shared the same sequence. Frame shifting, termination, deletion, and substitution accounted for the heterogeneity of RT sequences. Southern blot hybridization using the RT1 clone from the “Cara Cara” navel as a probe showed that multiple copies were integrated throughout the sweet orange genomes, which made the retrotransposon possible an effective molecular marker to detect citrus evolution events and to reveal its relationship with bud mutation. No transcriptional activities of the retrotransposon were detected by RT-PCR and Northern analysis in the fruits and leaves of either “Cara Cara” or “Seike” navels.

Effective Isolation of Retrotransposons and Repetitive DNA Families from the Wheat Genome

New classes of repetitive DNA elements were effectively identified by isolating small fragments of the elements from the wheat genome. A wheat A genome library was constructed from Triticum monococcum by degenerate cleavage with EcoO1091, the recognition sites of which consisted of 5＇-PuGGNCCPy-3＇ multi-sequences. Three novel repetitive sequences pTm6, pTm69 and pTm58 derived from the A genome were screened and tested for high copy number using a blotting approach, pTm6 showed identity with integrase domains of the barley Tyl-Copia-retrotransposon BARE-1 and pTm58 showed similarity to the barley Ty3-gypsy-like retrotransposon Romani. pTm69, however, constituted a tandem array with useful genomic specificities, but did not share any identity with known repetitive elements. This study also sought to isolate wheat D-genome-specific repetitive elements regardless of the level of methylation, by genomic subtraction. Total genomic DNA of Aegilops tauschii was cleaved into short fragments with a methylation-insensitive 4 bp cutter, Mbol, and then common DNA sequences between Ae. tauschii and Triticum turgidum were subtracted by annealing with excess T. turgidum genomic DNA. The D genome repetitive sequence pAt1 was isolated and used to identify an additional novel repetitive sequence family from wheat bacterial artificial chromosomes with a size range of 1 395-1 850 bp. The methods successfully led pathfinding of two unique repetitive families.

Genome Size Evolution Mediated by Gypsy Retrotransposons in Brassicaceae

The dynamic activity of transposable elements(TEs)contributes to the vast diversity of genome size and architecture among plants.Here,we examined the genomic distribution and transposition activity of long terminal repeat retrotransposons(LTR-RTs)in Arabidopsis thaliana(Ath)and three of its relatives,Arabidopsis lyrata(Aly),Eutrema salsugineum(Esa),and Schrenkiella parvula(Spa),in Brassicaceae.Our analyses revealed the distinct evolutionary dynamics of Gypsy retrotransposons,which reflects the different patterns of genome size changes of the four species over the past million years.The rate of Gypsy transposition in Aly is approximately five times more rapid than that of Ath and Esa,suggesting an expanding Aly genome.Gypsy insertions in Esa are strictly confined to pericentromeric heterochromatin and associated with dramatic centromere expansion.In contrast,Gypsy insertions in Spa have been largely suppressed over the last million years,likely as a result of a combination of an inherent molecular mechanism of preferential DNA removal and purifying selection at Gypsy elements.Additionally,species-specific clades of Gypsy elements shaped the distinct genome architectures of Aly and Esa.

New Insights into Nested Long Terminal Repeat Retrotransposons in Brassica Species

Long terminal repeat （LTR） retrotransposons, one of the foremost types of transposons, continually change or modify gene function and reorganize the genome through bursts of dramatic proliferation. Many LTR-TEs preferen-tially insert within other LTR-TEs, but the cause and evolutionary significance of these nested LTR-TEs are not well under-stood. In this study, a total of 1.52 Gb of Brassica sequence containing 2020 bacterial artificial chromosomes （BACs） was scanned, and six bacterial artificial chromosome （BAC） clones with extremely nested LTR-TEs （LTR-TEs density： 7.24/kb） were selected for further analysis. The majority of the LTR-TEs in four of the six BACs were found to be derived from the rapid proliferation of retrotransposons originating within the BAC regions, with only a few LTR-TEs originating from the proliferation and insertion of retrotransposons from outside the BAC regions approximately 5-23 Mya. LTR-TEs also pref-erably inserted into TA-rich repeat regions. Gene prediction by Genescan identified 207 genes in the 0.84Mb of total BAC sequences. Only a few genes （3/207） could be matched to the Brassica expressed sequence tag （EST） database, indicating that most genes were inactive after retrotransposon insertion. Five of the six BACs were putatively centromeric. Hence, nested LTR-TEs in centromere regions are rapidly duplicated, repeatedly inserted, and act to suppress activity of genes and to reshuffle the structure of the centromeric sequences. Our results suggest that LTR-TEs burst and proliferate on a local scale to create nested LTR-TE regions, and that these nested LTR-TEs play a role in the formation of centromeres.

Genome-wide Characterization of Long Terminal Repeat-retrotransposons in Apple Reveals the Differences in Heterogeneity and Copy Number between Ty1-copia and Ty3-gypsy Retrotransposons

The conserved domains of reverse transcriptase （RT） genes of Tyl-copia and Ty3-gypsy groups of long terminal repeat （LTR） retrotransposons were isolated from the Malus domestica genome using degenerate oligonucleotide primers. Sequence analysis showed that 45% of Ty1-copia and 63% of Ty3-gypsy RT sequences contained premature stop codons and/or indels disrupting the reading frame. High heterogeneity among RT sequences of both Ty1-copia and Ty3-gypsy group retrotransposons was observed, but Ty3-gypsy group retrotransposons in the apple genome are less heterogeneous than Ty1-copia elements. Retrotransposon copy number was estimated by dot blot hybridizations for Ty1-copia （-5000） and Ty3-gypsy （-26000）. All elements of the two types of LTR retrotransposons comprise approximately 38% of the M. domestica genome, with the Ty3-gypsy group contribution being higher （33.5%） than the Tyl.copia one （4.6%）. Transcription was not detected by reverse transcription-polymerase chain reaction for either Ty1-copia or Ty3-gypsy retrotransposons in the leaves of plants in vitro or in leaf explants cultured on medium supplemented with high concentration benzylaminopurine. This research reveals the differences in heterogeneity and copy number between Ty1-copia and Ty3-gypsy retrotransposons in the apple genome. Ty1-copia retrotransposon has higher heterogeneity than Ty3-gypsy retrotransposon, but the latter has a higher copy number, which implies that Ty3-gypsy retrotransposons may play a more important role in the apple genome evolution.

Natural selection maintains the transcribed LTR retrotransposons in Nosema bombycis

Eight intact LTR retrotransposons （Nbr1-Nbr8） have been previously characterized from the genome of Nosema bombycis, a eu- karyotic parasite with a compact and reduced genome. Here we describe six novel transcribed Nbr elements （Nbr9-Nbr14） identified through either cDNA library or RT-PCR. Like previously determined ones, all of them belong to the Ty3/Gypsy superfamily. Retrotransposon diversity and incomplete domains with insertions （Nbr12）, deletions （Nbrll） and in-frame stop codons in coding regions （Nbr9） were detected, suggesting that both defective and loss events of LTR retrotransposon have happened in N. bornbycis genome. Analysis of selection showed that strong purifying selection acts on all elements except Nbr11. This implies that selective pressure keeps both these Nbrs and their functions in genome. Interestingly, Nbrll is under positive selection and some positively selected codons were identified, indicating that new functionality might have evolved in the Nbrll retrotransposon. Unlike other transposable elements, Nbrll has integrated into a conserved syntenic block and probably resulted in the inversion of both flanking regions. This demonstrates that transposable element is an important factor for the reshuffling and evolution of their host genomes, and may be maintained under natural selection.

Genomic Rearrangement in Endogenous Long Terminal Repeat Retrotransposons of Rice Lines Introgressed by Wild Rice （Zizania latifolia Griseb.）

Stochastic introgression of alien DNA may impose a genomic stress to the recipient genome. Herein, we report that apparent de novo genomic rearrangements in 10 of 13 selected endogenous, lowcopy, and potentially active long terminal repeat （LTR） retrotransposons occurred in one or more of three rice lines studied that were introgressed by wild rice （Zizania latifolia Griseb.）. For nine retrotransposons in which both the reverse-transcriptase （RT） region and the LTR region were available, largely concordant rearrangements occurred at both regions in five elements and at the RT region only in the remaining four elements. A marked proportion of the genomic changes was shared by two or all three introgression lines that were derived from a single F~ plant. This indicates that most of the genomic changes occurred at early developmental stages of the F~ somatic cells, which then gave rise to germline cells, and, hence, ensured inheritance of the changes to later generations. Possible causes and potential implications of the introgression-induced genomic rearrangements in LTR retrotransposons are discussed in the context of plant genome evolution and breeding.

Retrotransposons in pluripotent stem cells

Transposable elements constitute about half of the mammalian genome,and can be divided into two classes:the class I(retrotransposons)and the class II(DNA transposons).A few hundred types of retrotransposons,which are dynamic and stage specific,have been annotated.The copy numbers and genomic locations are significantly varied in species.Retrotransposons are active in germ cells,early embryos and pluripotent stem cells(PSCs)correlated with low levels of DNA methylation in epigenetic regulation.Some key pluripotency transcriptional factors(such as OCT4,SOX2,and NANOG)bind retrotransposons and regulate their activities in PSCs,suggesting a vital role of retrotransposons in pluripotency maintenance and self-renewal.In response to retrotransposons transposition,cells employ a number of silencing mechanisms,such as DNA methylation and histone modification.This review summarizes expression patterns,functions,and regulation of retrotransposons in PSCs and early embryonic development.

A 314-bp SINE insertion in the ZNF2 promoter region may act as a repressor related to regulation of fat deposition in pigs

Retrotransposons,a type of DNA fragment that can mobilize itself on genome,can generate genetic variations and develop for molecular markers based on the insertion polymorphism.Zinc finger proteins(ZNFs)are among the most abundant proteins in eukaryotic animals,and their functions are extraordinarily diverse and particularly important in gene regulation.In the current study,bioinformatic prediction was performed to screen for retrotransposon insertion polymorphisms(RIPs)in six ZNF genes(ZNF2,ZNF3,ZNF7,ZNF8,ZNF10 and ZNF12).Six RIPs in these ZNFs,including one short interspersed nuclear element(SINE)RIP in intron 1 and one long interspersed nuclear element 1(L1)RIP in intron 3 of ZNF2,one SINE RIP in 5′flanking region and one SINE RIP in intron 2 of ZNF3,one SINE RIP in 3′UTR of ZNF7 and one L1 RIP in intron 2 of ZNF12,were discovered and their presence was confirmed by PCR.The impact of the SINE RIP in the first intron of ZNF2,which is close to the core promoter of ZNF2,on the gene activity was investigated by dual-luciferase assay in three cell lines.Our results showed that the SINE insertion in the intron 1 of ZNF2 repressed the core promoter activity extremely significantly(P<0.01)in cervical cancer cells and porcine primary embryonic fibroblasts(HeLa and PEF),thus SINE may act as a repressor.This SINE RIP also significantly(P<0.05)affected the corrected back fat thickness in Yorkshire pigs.The corrected back fat thickness of individuals with SINE insertion in the first intron of ZNF2 was significantly(P<0.05)higher than that of individuals without SINE insertion.In summary,our data suggested that RIPs play important roles in the genetic variations of these ZNF genes and SINE RIP in the intron 1 of ZNF2 may provide a useful molecular marker for the screening of fat deposition in the pig breeding.

Identification and Characterization of Reverse Transcriptase Fragments of Long Interspersed Nuclear Elements (LINEs) in the <i>Morus notabilis</i>Genome

Reverse transcriptase (rt) fragments from LINE retrotransposons in the mulberry genome were analyzed in terms of heterogeneity, phylogeny, and chromosomal distribution. We amplified and characterized conserved domains of the rt using degenerate primer pairs. Sequence analyses indicated that the rt fragments were highly heterogeneous and rich in A/T bases. The sequence identity ranged from 31.8% to 99.4%. Based on sequence similarities, the rt fragments were categorized into eight groups. Furthermore, similar stop codon distribution patterns among a series of clones in the same group indicated that they underwent a similar evolutionary process. Interestingly, phylogenetic analyses of the rt fragments isolated from mulberry and 13 other plant species revealed that two distantly related taxa (mulberry and Paeonia suffruticosa) grouped together. It does not appear that this phenomenon resulted from horizontal transposable element transfer. Fluorescence in situ hybridization analysis revealed that most of the rt fragments were concentrated in the subtelomeric and pericentromeric regions of the mulberry chromosomes, but that these elements were not abundant in the mulberry genome. Future studies will focus on the potential roles of these elements in the subtelomeric and pericentromeric regions of the mulberry genome.

<i>At</i>L1 a Non-LTR Retrotrasposon Fragment in the Genome of <i>Arabidopsis thaliana </i>with Homology to Plants and Animals

We report the isolation of AtL1, a 249 bp non-LTR retrotransposon fragment from Arabidopsis thaliana by fingerprinting mRNAs extracted from A. thaliana plants, ecotype Columbia, in different heat stress conditions. Southern blot and PCR analysis suggested that AtL1 occurs as a single- or low-copy insert in the genome of A. thaliana ecotype Columbia. The presence of AtL1 in the genome of different Arabidopsis ecotypes was confirmed by PCR amplification and sequencing thus excluding all possible contamination. A preliminary scan of the AtL1 nucleotide sequence against the EMBL and NCBI databases revealed a high degree of similarity to a group of LINE type L1 retrotransposons of mammals and with a cDNA sequence of Artemisia annua. A phylogenetic analysis of LINE elements from animals and plants placed AtL1 and A. annua sequences in close proximity to some mammalian sequences but distant from the other plants LINE elements including those from Arabidopsis.

Renal cell carcinoma and viral infections:A dangerous relationship?

Virus-related cancers in humans are widely recognized,but in the case of renal cancer,the link with the world of viruses is not clearly established in humans,despite being known in animal biology.In the present review,we aimed to explore the literature on renal cell carcinoma(RCC)for a possible role of viruses in human RCC tumorigenesis and immune homeostasis,hypothesizing the contribution of viruses to the immunogenicity of this tumor.A scientific literature search was conducted using the PubMed,Web of Science,and Google Scholar databases with the keywords“virus”or“viruses”or“viral infection”matched with(“AND”)“renal cell carcinoma”or“kidney cancer”or“renal cancer”or“renal carcinoma”or“renal tumor”or“RCC”.The retrieved findings evidenced two main aspects testifying to the relationship between RCC and viruses:The presence of viruses within the tumor,especially in non-clear cell RCC cases,and RCC occurrence in cases with pre-existing chronic viral infections.Some retrieved translational and clinical data suggest the possible contribution of viruses,particularly Epstein-Barr virus,to the marked immunogenicity of sarcomatoid RCC.In addition,it was revealed the possible role of endogenous retrovirus reactivation in RCC oncogenesis,introducing new fascinating hypotheses about this tumor’s immunogenicity and likeliness of response to immune checkpoint inhibitors.

Retrotransposon-mediated DELLA transcriptional reprograming underlies semi-dominant dwarfism in foxtail millet

Retrotransposons account for a large proportion of the genome and genomic variation, and play key roles in creating novel genes and diversifying the genome in many eukaryotic species. Although retrotransposons are abundant in plants, their roles had been underestimated because of a lack of research. Here, we characterized a gibberellin Acid (GA)-insensitive dwarf mutant, 84133, in foxtail millet. Map-based cloning revealed a 5.5-kb Copia-like retrotransposon insertion in DWARF1 (D1), which encodes a DELLA protein. Transcriptional analysis showed that the Copia retrotransposon mediated the transcriptional reprogramming of D1 leading to a novel N-terminal-deleted truncated DELLA transcript that was putatively driven by Copia's LTR, namely D1-TT, and another chimeric transcript. The presence of D1-TT was confirmed by protein immunodetection analysis. Furthermore, D1-TT protein was resistant to GA3 treatment compared with the intact DELLA protein due to its inability to interact with the GA receptor, SiGID1. Overexpression of D1-TT in foxtail millet resulted in dwarf plants, confirming that it determines the dwarfism of 84133. Thus, our study documents a rare instance of long terminal repeat (LTR) retrotransposon-mediated transcriptional reprograming in the plant kingdom. These results shed light on the function of LTR retrotransposons in generating new gene functions and genetic diversity.

VL30 retrotransposition signals activation of a caspaseindependent and p53-dependent death pathway associated with mitochondrial and lysosomal damage

长终端重复(LTR ) 的影响房间命运上的 retrotransposition 是未知的。这里，我们在转变 SV40 的老鼠 SVTT1 房间在房间死亡上调查了 VL30 retrotransposition 的效果。VL30 retrotransposon 的 Transfection 减少了由 17 褶层的 SVTT1 的 clonogenicity，作为与父母 NIH3T3 房间相比。频率和房间死亡率在 retrotransposition 积极的 SVTT1 被发现的 retrotransposition 的相关层次克隆房间，展出 DNA 破碎，原子冷凝作用， multinucleation 和细胞质的 vacuolization。受动器 caspases 的激活的分析揭示了 caspase 独立的房间死亡机制。然而，房间死亡与感应的 p53 和 Bcl-2 和 Hsp70 蛋白质表示的美洲狮伪和 Bax 和 downregulation 的伴随物 upregulation 被联系。而且，我们发现了大 T 抗原(副)/p53 和 p53 translocation 的 colocalization 的部分损失到线粒体，导致外部膜 permeabilization (MOMP ) 由 lysosomal 膜 permeabilization (LMP ) 伴随了的 mitochondrial。有趣地，有抗氧化剂 N-acetylcysteine 的处理废除了细胞死亡，建议导出 mitochondrial 的反应的氧种类的参与，并且导致了 retrotransposition 频率的增加。重要地，当 VL30 动员被导致，房间死亡的正式就职是 retrotransposon 特定的 VL30；相反， non-LTR L1 的动员(LINE-1，长散布的原子 element-1 ) ， B2 和 LTR MusD retrotransposons 减少了。第一次，我们的结果提供 VL30 retrotransposition 调停的充分证据经由 mitochondrial 和 lysosomal 损坏的房间死亡，揭开是的 retrotransposition 的角色在触发房间死亡激活 mitochondrial-lysosomal 串音的一个原子信号。

Characterization of a <i>Tos</i>17 Insertion Mutant of Rice Auxin Signal Transcription Factor Gene, <i>OsARF</i>24

Auxin signaling plays a key role in the regulation of various growth and developmental processes in higher plants. Auxin response factors (ARFs) are transcription factors that regulate the expression of auxin-response genes. The osarf24-1 mutant contains a truncation of domain IV in the C-terminal dimerization domain of a rice ARF protein, OsARF24. This mutant showed auxin-deficient phenotypes and reduced sensitivity to auxin. However, OsARF24 protein contains an SPL-rich repression domain in its middle region and acts as a transcriptional repressor. These results imply that the C-terminal dimerization domain, especially the C-terminal half of domain IV, is essential for the proper regulation of OsARF24 function as a transcriptional repressor in rice.

Recent amplification of Osr4 LTR-retrotransposon caused rice D1 gene mutation and dwarf phenotype

A novel rice d1 mutant was identified using map-based cloning and comparative analysis of known dl mutants.The mutant[d1-a) shows a mild dwarf trait,which differs only slightly from the wildtype in plant height at the tillering stage.The dl-a mutant is different from other dl mutants.We found that it was interrupted by an Osr4 long terminal repeat(LTR)-retrotransposon,which resulted in the loss of exon 7 in the mutant D1 mRNA.A paralog of the D1 gene.D1-like,was revealed.D1-like is a truncated gene that might have resulted from recombination between retrotransposons.We identified 65 Osr4LTR-retrotransposons in Nipponbare,and found more LTR variants in contrast to coding DNA sequence(CDS) in the retrotransposons.We also identified five possible regulatory motifs in LTRs which may control the expression of the retrotransposons.In addition,we predicted six putative functional Osr4 retrotransposons that contain complete CDSs and all important elements.Osr4 retrotransposons were classified into 4 groups,and this type of retrotransposon only appears to be present in monocots.Members of group 1-1,which included all putative functional retrotransposons,showed a high similarity with each other.The retrotransposons were expressed in all tissues,at especially higher levels in some leaves and seeds.These findings imply that transpositions of group 1-1 members might have occurred frequently and recently.

Differential Expression of Retrotransposon WIS 2-1A Response to Vacuum, Low-Energy N^+ Implantation and ^(60)Coγ-ray Irradiation in Wheat

Mutagenesis and retrotransposons have a close relationship, but little attention has been paid yet to the activity of retrotransposons produced by physical mutagens. The variation of retrotransposon WIS 2-1A activity in wheat （Triticum aestivum L.） embryos at three different growth times （30 h, 45 h and 60 h） was investigated after they had been treated with N^＋ implantation in a vacuum of 5× 10^-2 Pa and irradiation by ^60Coγ-ray respectively. For each of the three growth times the expression of WIS 2-1A showed almost entirely a same trend of downregulation, upregulation, then downregulation, and upregulation again with the increase in dose of N^＋ implantation, but the expression appeared irregular with the increase in irradiation of ^60Coγ-ray. In conclusion, the acutely activating effect of WIS 2-1A stimulated by vacuum and high dose N^＋ implantation within a shorter incubation time may provide a convenient tool to advance the research on mutagenic breeding and function genes.

Evaluation of Transgenic <i>Nicotiana tabacum</i>with <i>deh</i>E Gene Using Transposon Based IRAP Markers

In the present study, five genetically modified herbicide tolerant Nicotiana tabacum cv. TAPM24 plants with a constructed vector pCAMBIA1301a carrying dehalogenase E (dehE) gene were compared with three non-transgenic controls using Tto1 retrotransposon specific IRAP markers. dehE gene was originally characterized in Rhizobium sp. and it produced an enzyme which degraded the Dalapon herbicide. IRAP protocol was applied on transgenic and non-transgenic plants to investigate the retrotransposon based genetic variation which may appear during transformation. Polymorphism rates were calculated as 0%-20% from IRAP-PCR products among all plant samples. These results show that transformation of tobacco plant with the dehE gene may cause Tto1 retrotransposon alterations appearing as different band profiles. The findings are expected to contribute to genetic engineering studies to obtain better results and also to understand how transposons contribute to features such as transgenesis. In our knowledge, this is one of the first experimental data of transgenic N. tabacum engineered with dehE gene originated Rhizobium sp. in terms of retrotranposon based variation.