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The current data clearly show that H4-12 3′-UTR binds the SLBP and undergoes dynamic structural rearrangements that favour U7 snRNA hybridization. Such reaction is reminiscent of U6 snRNA which displays a very compact structure in its naked form, whereas in presence of Prp24p and Lsm proteins the RNA structure is much ... | 16982637_p22 | 16982637 | DISCUSSION | 4.455654 | biomedical | Study | [
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An interesting consequence of the SLBP role in promoting U7 binding is that 3′ end processing reaction of histone pre-mRNAs is an ordered process. The SLBP binds first, facilitates anchoring of U7 snRNP and finally the whole complex is locked by the ZFP100, which bridges the SLBP-hairpin complex to the U7 snRNP. This m... | 16982637_p23 | 16982637 | DISCUSSION | 4.636237 | biomedical | Study | [
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To conclude, the SLBP is necessary for 3′ end processing of histone pre-mRNAs. In addition, the SLBP is essential for cell-cycle regulation of histone expression. The SLBP is one of the three actually known cell cycle regulated factors that are involved in the 3′ end processing reaction of the histone pre-mRNAs ( 32 , ... | 16982637_p24 | 16982637 | DISCUSSION | 4.288258 | biomedical | Study | [
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Retrotransposition is a wide spread phenomenon occurring in eukaryotic genomes of diverse taxonomic groups. It is believed to be responsible for various important events in the genome, such as gene inactivation, transduction of genomic sequences, regulation of gene expression and genome expansion ( 1 ). It has also bee... | 17040894_p0 | 17040894 | INTRODUCTION | 4.117502 | biomedical | Study | [
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Entamoeba histolytica , a primitive eukaryote, is the third leading cause of morbidity and mortality due to parasitic disease in humans, and is estimated to be responsible for between 50 000 and 100 000 deaths every year ( 3 ). It is home to the non-LTR retrotransposons EhLINEs and EhSINEs. These together account for a... | 17040894_p1 | 17040894 | INTRODUCTION | 4.347541 | biomedical | Study | [
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0.9986478686332703,
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Target primed reverse transcription (TPRT) is thought to be the mechanism by which non-LTR retrotransposons insert in the genome ( 6 ). Since retrotransposition is initiated by the element-encoded endonuclease (EN) making a nick at the bottom strand of the site of insertion, an important determinant of target site spec... | 17040894_p2 | 17040894 | INTRODUCTION | 4.627788 | biomedical | Study | [
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0.9968343377113342,
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The apparent lack of targeted insertion of many non-LTR elements could be due to non sequence specific nicking by the element-encoded EN, or it may imply that these elements recognize structural features of the DNA rather than sequence alone. Do the insertion sites share conserved structural features which are recogniz... | 17040894_p3 | 17040894 | INTRODUCTION | 4.476753 | biomedical | Study | [
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] | [
0.9988991022109985,
0.0002859982487279922,
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] | en | 0.999996 |
EhLINE1-EN protein was purified as described ( 22 ) except that Escherichia coli cells were grown for 90 min after adding isopropyl-β- d -thiogalactopyranoside (IPTG). The recombinant protein was eluted with 250 mM imidazole after extensive washing with buffer containing 80 mM imidazole. The protein was immediately dia... | 17040894_p4 | 17040894 | Expression and purification of EhLINE1-EN | 4.134273 | biomedical | Study | [
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] | [
0.9951019287109375,
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] | en | 0.999996 |
For preparation of radiolabeled substrates by PCR, the bottom strand primer (50 pmol) was end labeled in a 20 μl reaction using 50 μCi of [γ- 32 P]ATP (Amersham pharmacia Biosciences) and T4 polynucleotide kinase (NEB). The reaction was stopped by incubating at 65°C for 20 min and the labeled primer was purified by pas... | 17040894_p5 | 17040894 | Preparation of substrates and EN assays | 4.158287 | biomedical | Study | [
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] | en | 0.999996 |
The DNA substrate (100 ng) was incubated with 40 ng EN protein in a 10 μl reaction for 1 h at 37°C. The enzyme was inactivated by adding 25 mM EDTA. Denaturing electrophoresis was performed on 6–12% polyacrylamide gels containing 7 M urea. A 2 μl aliquot of the reaction product was mixed with 8 μl of formamide gel load... | 17040894_p6 | 17040894 | Preparation of substrates and EN assays | 4.190656 | biomedical | Study | [
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] | [
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] | en | 0.999997 |
Synthetic substrates (32–35 bp) and the 27 bp substrate were prepared by annealing the overlapping complementary single-stranded oligonucleotides, followed by gap filling and PCR. The substrates were purified and treated with EN as described above. | 17040894_p7 | 17040894 | Preparation of substrates and EN assays | 3.955081 | biomedical | Study | [
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The insertion loci were obtained by using an automated software tool ELEANALYSER that was developed for analysis of elements in a genome ( 4 ). This tool incorporates various Perl programs as filters and parsers along with BLAST ( 24 ) suite of programs. The target site duplications at the boundaries of elements were d... | 17040894_p8 | 17040894 | Data retrieval | 3.994383 | biomedical | Study | [
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] | en | 0.999998 |
The positive dataset consisted of 93 sequences of known insertion sites (see the Results section) while the negative dataset consisted of 100 sequences known not to permit insertion. For each of the structural properties discussed here, a graphical profile was constructed for each member of the positive or negative dat... | 17040894_p9 | 17040894 | Computational analysis of E.histolytica pre insertion sequences | 4.115128 | biomedical | Study | [
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] | en | 0.999996 |
In order to measure if differences between the controls and the insertion sites were significant, we used Mann–Whitney tests using the statistical software MINITAB. Analysis of any DNA sequence with respect to the different parameters described here is possible at this website with the tool DNA SCANNER. | 17040894_p10 | 17040894 | Computational analysis of E.histolytica pre insertion sequences | 2.786576 | biomedical | Study | [
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] | en | 0.999997 |
In a similar manner, we constructed positive and negative datasets for other genomes. Namely, upstream regions for a set of known insertion sites were curated from GenBank for the organisms, such as Dictyostelium discoideum, Takifugu rubripes and Drosophila melanogaster . A negative dataset was also constructed as foll... | 17040894_p11 | 17040894 | Insertion site analysis in other genomes | 4.075225 | biomedical | Study | [
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] | [
0.9994114637374878,
0.00020147541363257915,
0.00034528030664660037,
0.00004185895886621438
] | en | 0.999999 |
According to the TPRT model of retrotransposition by non-LTR elements, the process is initiated by a nick in the bottom strand of the target site, generated by EN. An important determinant in the choice of insertion sites by EhLINEs/SINEs could, therefore, be the preferred substrate requirements of the EN. To test this... | 17040894_p12 | 17040894 | Role of the element-encoded EN in target site selection | 4.316969 | biomedical | Study | [
0.9993239641189575,
0.0003588764520827681,
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] | [
0.9993927478790283,
0.00023472662724088877,
0.0002950765483547002,
0.00007746947085252032
] | en | 0.999995 |
It had earlier been observed that a G residue was frequently present 3–4 nt upstream of the nick in most sites nicked by the EN ( 22 ). This may be significant given that the E.histolytica genome is highly A + T rich. ( 26 ). Site #3 contains three G residues upstream of the nick . Using the procedure described above, ... | 17040894_p13 | 17040894 | Role of the element-encoded EN in target site selection | 4.221298 | biomedical | Study | [
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0.00028709316393360496,
0.0003146679373458028
] | [
0.9995085000991821,
0.00019244947179686278,
0.0002326836111024022,
0.00006632513395743445
] | en | 0.999996 |
To validate the general applicability of this sequence requirement, site #2 in the 176 bp fragment was also analyzed by mutation analysis. A fragment of 85 bp (position 92 to 176) was PCR amplified from the 176 bp template. The ‘wild-type’ sequence of this site (bottom strand) was 5′-TGCATTG-3′. In agreement with the r... | 17040894_p14 | 17040894 | Role of the element-encoded EN in target site selection | 4.152061 | biomedical | Study | [
0.999417781829834,
0.0002645309432409704,
0.00031769939232617617
] | [
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0.00026639472343958914,
0.0001719904539640993,
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] | en | 0.999998 |
Nucleotides in the vicinity of the nicking site were checked for their role in substrate recognition. A 37 bp substrate containing 15 bp upstream and 22 bp downstream of the nick in site #3 was used. Transition mutations were introduced in every alternate nucleotide, keeping the central 9 bp (GAATACCTC) unchanged . Thi... | 17040894_p15 | 17040894 | Role of the element-encoded EN in target site selection | 4.122827 | biomedical | Study | [
0.9993526339530945,
0.0003082608454860747,
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] | [
0.9994572997093201,
0.0002902369888033718,
0.00018685428949538618,
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] | en | 0.999997 |
The above substrates were derived from a natural E.histolytica sequence in which EhSINE1 is known to insert. A completely artificial substrate was next tested for enzyme activity with EN. It was made AT-rich and variants containing two Cs or two Gs were also tested . The results confirmed the earlier observations with ... | 17040894_p16 | 17040894 | Role of the element-encoded EN in target site selection | 4.156594 | biomedical | Study | [
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] | [
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0.00027204849175177515,
0.0002554336388129741,
0.00005815616168547422
] | en | 0.999996 |
The above data shows that EhLINE1-encoded EN, while being flexible in its sequence requirement, has a strong preference for nicking the bottom strand between A and T residues located downstream of GC (5′-GCATT-3′). Sequences further upstream or downstream of this basic sequence had little or no effect on enzyme activit... | 17040894_p17 | 17040894 | Role of the element-encoded EN in target site selection | 4.098845 | biomedical | Study | [
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0.0002266072406200692,
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] | [
0.9988839030265808,
0.0008465697756037116,
0.00018936801643576473,
0.0000802424328867346
] | en | 0.999996 |
The 176 bp fragment of E.histolytica DNA used as a substrate for EN in the above experiments has three hotspots of nicking by EN. Of these, EhSINE1 is known to insert at site #3. We tested whether site #2, which was also efficiently nicked by the EN, was used as an integration site for these elements. Primers were desi... | 17040894_p18 | 17040894 | The preferred EN recognition sequence alone is insufficient for element insertion | 4.261924 | biomedical | Study | [
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] | [
0.9994843006134033,
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0.00007258751429617405
] | en | 0.999997 |
We selected all the EhSINE1 insertion sites in which the 5′ end of EhSINE1 could be clearly identified. These numbered a total of 93 and were used to construct a set of pre insertion loci of EhSINE1 as follows. Each occupied EhSINE1 site was analyzed and the element, together with one of the target site duplications, w... | 17040894_p19 | 17040894 | Structural features of the insertion site of EhSINE1 as deduced from computational analysis | 4.130003 | biomedical | Study | [
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] | [
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0.0002183698961744085,
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] | en | 0.999997 |
Positive and negative datasets were compared with respect to the following criteria: DNA sequence, structure, energy profiles, protein induced deformability and nucleosome location. Computation of nine measures was performed in a moving window of length 5 over each 80 bp segment, and the profile was averaged for all lo... | 17040894_p20 | 17040894 | Structural features of the insertion site of EhSINE1 as deduced from computational analysis | 4.377586 | biomedical | Study | [
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] | en | 0.999997 |
When the 93 true insertion sites were tested with the nine measures listed above, for 10 of these sites none of the measures scored positive . For the remaining 83 sites one or more of the measures scored positive, with more than half the sites scoring positive on four or more of the measures. The 5754 E.histolytica ge... | 17040894_p21 | 17040894 | Structural features of the insertion site of EhSINE1 as deduced from computational analysis | 4.111111 | biomedical | Study | [
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0.9995583891868591,
0.0002073518407996744,
0.00018527291831560433,
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] | en | 0.999998 |
Since, DNA structure at pre insertion loci of EhSINE1 was distinct, we further checked to see if structure had an influence on EhLINE1-encoded EN activity as well. The various mutated substrates used to check EN activity were analyzed for changes in DNA structure as a result of the introduced mutations. The substrates ... | 17040894_p22 | 17040894 | Computational analysis of DNA structure adopted by mutated substrates of the EhLINE1-EN | 4.120269 | biomedical | Study | [
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] | [
0.9996216297149658,
0.0001322414173046127,
0.0001985888957278803,
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] | en | 0.999997 |
To see if the physical features listed above for EhSINE1 insertion sites were shared by retrotransposon insertion sites in other genomes as well, a few selected genomes were analyzed using DNA SCANNER ( Table 2 ). Site-specific as well as non site-specific elements were analyzed in each genome. A stretch of 40 bp upstr... | 17040894_p23 | 17040894 | Insertion sites of many non-LTR retrotransposons share common structural features | 4.147452 | biomedical | Study | [
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] | [
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] | en | 0.999995 |
While all the measures used in the present study to detect the insertion sites of elements of E.histolytica are not universally applicable in other genomes, the present observations suggest the possibility of subsets of these properties being pertinent for different organisms. Although, the structure and intensity of t... | 17040894_p24 | 17040894 | Insertion sites of many non-LTR retrotransposons share common structural features | 4.029667 | biomedical | Study | [
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0.0004809668753296137,
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] | en | 0.999996 |
Amongst parasitic protozoa E.histolytica is one of the few in which non-LTR retrotransposons occupy as much as 6–8% of its 23 Mb genome ( 4 , 34 ). From a phylogenetic standpoint it is important to understand whether this primitive organism shares the same mechanisms for insertion and maintenance of these elements in i... | 17040894_p25 | 17040894 | DISCUSSION | 4.173186 | biomedical | Study | [
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Different parameters that probed structural, thermodynamic or nucleosome positioning features were employed in our computational analysis of target site sequences in order to detect unique features, which may be recognized by the invading retrotransposon ( Table 2 ). This analysis showed that DNA structure is likely to... | 17040894_p26 | 17040894 | DISCUSSION | 4.205297 | biomedical | Study | [
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In our analysis of E.histolytica the most significant outcome was that in all insertion sites of EhSINE1 the region −10 to −35 bp upstream of the insertion point showed a very distinct structure. This region was also T-rich. However, the observed profiles were not attributable to T-richness alone, since shuffling the s... | 17040894_p27 | 17040894 | DISCUSSION | 4.282293 | biomedical | Study | [
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0.9994638562202454,
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] | en | 0.999996 |
We have examined the sequence requirements of the EhLINE1-encoded EN and find that although the enzyme is not strictly sequence-specific (although belonging to the REL-ENDO class), it is possible to assign a consensus sequence 5′-GCATT-3′ at which the enzyme nicks most efficiently between A-T and T-T. The upstream G wa... | 17040894_p28 | 17040894 | DISCUSSION | 4.334926 | biomedical | Study | [
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] | [
0.9993554949760437,
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] | en | 0.999995 |
Although, the in vitro consensus sequences preferred by EN are widely distributed in the genome, both in genic as well as intergenic regions, EhLINE1/SINE1 insertions have not been found within any gene so far. The preference of EhLINE1/SINE1 for intergenic regions would minimize direct damage to genes by insertional i... | 17040894_p29 | 17040894 | DISCUSSION | 4.193258 | biomedical | Study | [
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0.9990690350532532,
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] | en | 0.999997 |
In our earlier model of EhLINE1/SINE1 retrotransposition we had proposed a melting of the DNA duplex in the T-rich upstream region to allow positioning of the element RNA by virtue of hydrogen bonding between its T-rich 3′-tail and the A-rich bottom strand of DNA ( 22 ). In this context it is significant that the same ... | 17040894_p30 | 17040894 | DISCUSSION | 4.544942 | biomedical | Study | [
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In summary, our combination of computational and enzymatic analysis of pre-insertion loci can lead to a more realistic understanding of why these genomic loci are preferred for retrotransposition. | 17040894_p31 | 17040894 | DISCUSSION | 3.845792 | biomedical | Study | [
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Skeletal muscle formation in vertebrates involves an interplay between the maturation of the contractile and the neural apparatus which is intricate and interdependent ( 1 – 3 ). Coordinated events in myogenesis and neuromuscular junction formation are facilitated by intercellular communication between myoprogenitors a... | 17062625_p0 | 17062625 | INTRODUCTION | 4.327832 | biomedical | Study | [
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Communication between developing muscle fibers is extensive during prenatal embryonic development. In fact, coupling is so extensive prenatally that excitation spreading laterally between myotubes gives rise to waves of excitation that propagate across the entire muscle ( 1 ). In developing intercostals muscles, for in... | 17062625_p1 | 17062625 | INTRODUCTION | 4.87368 | biomedical | Study | [
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] | [
0.9703370928764343,
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In vivo observations have largely agreed with studies done on isolated primary myoblast cultures and with myoblast cell lines. Cx43 is expressed in established myoblast cell lines, and in isolated primary myoblast cultures prior to fusion ( 11 , 12 ). The fusion of myoblasts in vitro has been shown to require the prese... | 17062625_p2 | 17062625 | INTRODUCTION | 4.287226 | biomedical | Study | [
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] | [
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] | en | 0.999997 |
In myoblast cell lines, Cx43 is present prior to and during fusion but is rapidly downregulated after induction of differentiation despite only a slight reduction in mRNA levels ( 6 ). These studies suggested that the initial step in downregulation of Cx43 occurs at the translational level. In this study we present evi... | 17062625_p3 | 17062625 | INTRODUCTION | 4.267865 | biomedical | Study | [
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] | en | 0.999998 |
Alignment of DNA sequence pairs was performed using Blast 2 Sequences available at the NCBI website ( ) ( 16 ). Multiple DNA sequence alignments were performed using ClustalX v1.8 ( 17 ). All genomic, expressed sequence tags (ESTs), and other nucleotide sequences were obtained from the NCBI online database via Genomic ... | 17062625_p4 | 17062625 | Bioinformatics | 4.009953 | biomedical | Study | [
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0.00007523809472331777
] | en | 0.999996 |
Plasmids designed to express full length Cx43 mRNA sequence were subcloned from cDNA clone B0C01C08 from ATCC into the Xba I and Kpn I sites within pcDNA3.1 (Invitrogen). Reporter plasmids were constructed by PCR-amplifying the 3′-untranslated region (3′-UTR) from the cDNA clone and inserting the product into pcDNA3.1 ... | 17062625_p5 | 17062625 | Plasmid construction | 4.280729 | biomedical | Study | [
0.9995161294937134,
0.00024575492716394365,
0.00023801079078111798
] | [
0.9987454414367676,
0.0007621000404469669,
0.00039643023046664894,
0.00009601177589502186
] | en | 0.999997 |
C2C12 cells were a generous gift from Dr Charles Murry at the University of Washington were cultured as described ( 12 ). HeLa cells were cultured and transfected as described ( 19 ). Luciferase assays were performed as described ( 19 ). Antisense inhibitors of miR-1 and miR-206 as well as negative control oligonucleot... | 17062625_p6 | 17062625 | Cell culture | 4.071218 | biomedical | Study | [
0.9996740818023682,
0.00016133865574374795,
0.00016456676530651748
] | [
0.9988304972648621,
0.0006902381428517401,
0.0004140003293287009,
0.0000652898452244699
] | en | 0.999997 |
Total RNA was isolated from tissues using Trizol (Invitrogen) and a Tekmar homogenizer. Total RNA was isolated from cell cultures using Trizol. Total RNA from developmental stages of neonatal mice were purchased from Zyagen. The radiolabeled antisense RNA probe targeting miR-206 was prepared by using the pSUPER 206pri ... | 17062625_p7 | 17062625 | RNase protection assay (RPA) | 4.103028 | biomedical | Study | [
0.9996694326400757,
0.00013902189675718546,
0.00019156518101226538
] | [
0.9983258843421936,
0.0012377981329336762,
0.0003668112913146615,
0.000069555921072606
] | en | 0.999995 |
Total protein extracts from developmental stages of neonatal mice were purchased from Zyagen. Protein extracts were prepared in RIPA buffer [1% NP-40, 1% Deoxycholate, 0.1% SDS, 500 mM Tris, 150 mM NaCl, 1 mM PMSF, 1× Protease Inhibitor Cocktail (Roche)]. 10 μg of total protein was separated on precast 12% Tris–HCl PAG... | 17062625_p8 | 17062625 | Western blots | 4.091385 | biomedical | Study | [
0.9996299743652344,
0.00013914440933149308,
0.00023077873629517853
] | [
0.9978121519088745,
0.0017394585302099586,
0.0003752750635612756,
0.0000730736501282081
] | en | 0.999996 |
Northern blots designed to detect Cx43, Luciferase and β-actin mRNA were performed on 20 μg of total RNA from transfected cells using the NorthernMax Gly Kit (Ambion). Northern blots designed to detect miR-206 and U6 RNA from transfected HeLa cells were performed on 20 μg of total RNA by separating RNA on precast 15% T... | 17062625_p9 | 17062625 | Northern blotting | 4.161045 | biomedical | Study | [
0.9995912909507751,
0.000216075248317793,
0.00019267183961346745
] | [
0.9978657364845276,
0.0016064262017607689,
0.0004361695609986782,
0.0000916000353754498
] | en | 0.999998 |
C2C12 cells cultured in 35 mm dishes (Nunc) were induced to differentiate by switching to differentiation medium at ∼70% confluency. Cells were fixed at various time points by incubation for 5 min in −10°C methanol and air dryed. Cells were washed with PBS, further permeabilized for 5 min in 0.1% Triton in PBS and bloc... | 17062625_p10 | 17062625 | Immunocytochemistry | 4.157217 | biomedical | Study | [
0.999332845211029,
0.00044166046427562833,
0.0002254614228149876
] | [
0.9883192777633667,
0.010618488304316998,
0.0008170836954377592,
0.00024520294391550124
] | en | 0.999996 |
Based on evidence from previous work suggesting that downregulation of Cx43 during myogenesis occurs through a post-transcriptional mechanism, we looked for the presence of potential cis elements for translational regulation in the Cx43 mRNA sequence. Numerous investigators have published algorithms to detect possible ... | 17062625_p11 | 17062625 | Bioinformatics | 4.194767 | biomedical | Study | [
0.99950110912323,
0.000293773045996204,
0.0002050531911663711
] | [
0.9993731379508972,
0.00018288155843038112,
0.00036666521918959916,
0.00007730381184956059
] | en | 0.999995 |
To determine the expression pattern of miR-206, RPAs were performed on 5 μg of total RNA from various tissues. As shown in Figure 2 , miR-206 is expressed predominantly in adult skeletal muscle; it could also be detected in skin, E11 whole embryo and lung, but not in heart muscle. The absence of miR-206 in heart is con... | 17062625_p12 | 17062625 | Tissue distribution of miR-206 | 4.1312 | biomedical | Study | [
0.9995143413543701,
0.0002251557307317853,
0.00026052125031128526
] | [
0.9995085000991821,
0.00022999623615760356,
0.00021118084259796888,
0.00005041849362896755
] | en | 0.999997 |
In order to test if miR-206 is indeed capable of regulating Cx43 protein expression in the context of its native mRNA sequence, we cloned the full-length cDNA corresponding to the entire Cx43 mRNA sequence from transcription start site to polyadenylation signal into pcDNA3.1 (pcDNA Cx43). Another plasmid designed to de... | 17062625_p13 | 17062625 | Downregulation of Cx43 in vitro | 4.348916 | biomedical | Study | [
0.9994292855262756,
0.0003750818723347038,
0.00019566081755328923
] | [
0.9991346001625061,
0.00030561009771190584,
0.00044886957039125264,
0.00011093254579463974
] | en | 0.999996 |
To test the specific regulation of Cx43 through the two predicted binding sites, we inserted the Cx43 3′-UTR sequence downstream of the firefly luciferase coding region. Mutants of the putative binding sites were then prepared . Known sequence similarity between miR-206 and miR-1 caused us to also consider miR-1 as a p... | 17062625_p14 | 17062625 | Downregulation of Cx43 in vitro | 4.185035 | biomedical | Study | [
0.9994204044342041,
0.0003235973126720637,
0.0002560356224421412
] | [
0.9993845224380493,
0.00016206008149310946,
0.0003867563500534743,
0.00006667202978860587
] | en | 0.999996 |
Since we determined that miR-206 and to a lesser extent miR-1 are capable of regulating Cx43 expression through direct binding to the 3′-UTR of its mRNA and that miR-206 is expressed in skeletal muscle, the next step was to look in vivo to see if miR-206 regulates Cx43 expression during myogenesis. We predicted that mi... | 17062625_p15 | 17062625 | Regulation of Cx43 during development | 4.32453 | biomedical | Study | [
0.9994710087776184,
0.00033304927637800574,
0.00019599880033638328
] | [
0.999196469783783,
0.00023739523021504283,
0.0004712493682745844,
0.00009495379345025867
] | en | 0.999998 |
To confirm and extend our hypothesis that miR-206 plays an inportanat role in muscle cell development, we studied the expression of miR-206 in myoblasts in vitro , using a highly myogenic subclone of the mouse cell line C2C12. These cells are propagated as undifferentiated, mononucleated myoblasts under high serum cond... | 17062625_p16 | 17062625 | Downregulation of Cx43 in C2C12 cells | 4.277475 | biomedical | Study | [
0.9994276165962219,
0.0003712760517373681,
0.0002010539174079895
] | [
0.9993201494216919,
0.00022205838467925787,
0.00036236332380212843,
0.00009545003558741882
] | en | 0.999997 |
By day 2, expression of Cx43 had become undetectable in the perinuclear region and between cells, whereas myogenin expression had increased significantly. At this time, there were some multinucleated myotubes and some mononucleated myoblasts, none of which expressed Cx43. At day 3, most of the mononucleated cells had d... | 17062625_p17 | 17062625 | Downregulation of Cx43 in C2C12 cells | 4.2327 | biomedical | Study | [
0.9996058344841003,
0.00023889953445177525,
0.00015527885989286005
] | [
0.9988920092582703,
0.0004495815373957157,
0.0005621342570520937,
0.00009624506492400542
] | en | 0.999996 |
In order to further validate the in vivo correlations, we investigated the expression of Cx43 protein and mRNA as well as miR-206 during C2C12 myoblast differentiation. These cells were cultured in growth media and then induced to differentiate. Extracts were prepared at the time of induction and every 24 h thereafter.... | 17062625_p18 | 17062625 | Downregulation of Cx43 in C2C12 cells | 4.210863 | biomedical | Study | [
0.99946528673172,
0.0003317470254842192,
0.0002030764298979193
] | [
0.9994149208068848,
0.00017291821131948382,
0.00033509498462080956,
0.00007700992136960849
] | en | 0.999998 |
Finally, to show that miR-206 and miR-1 are indeed responsible for the downregulation of Cx43 in C2C12 cells, the cells were transfected with chemically modified antisense oligonucleotide microRNA inhibitors either individually or in combination immediately prior to induction of differentiation. Extracts were harvested... | 17062625_p19 | 17062625 | Downregulation of Cx43 in C2C12 cells | 4.129283 | biomedical | Study | [
0.9994944334030151,
0.00028244280838407576,
0.00022307061590254307
] | [
0.9994827508926392,
0.00016611095634289086,
0.00029200181597843766,
0.00005907805825700052
] | en | 0.999997 |
Skeletal muscle development in vertebrates is an evolutionarily conserved process that involves myoblast fusion into multinucleated myotubes. Prior to and during myoblast fusion, and during the development of the motor neuron input system, muscle precursors are electrically coupled via gap junctions. In mammals, as wel... | 17062625_p20 | 17062625 | DISCUSSION | 4.278356 | biomedical | Study | [
0.9996371269226074,
0.00020602378936018795,
0.00015678966883569956
] | [
0.9988652467727661,
0.0003601267817430198,
0.000689079228322953,
0.00008559657726436853
] | en | 0.999998 |
The prediction that miR-206 and to a lesser degree miR-1 would bind to the Cx43 mRNA by Lewis et al . ( 20 ) was one of many computer algorithms that predicted that a microRNA binding site was present in the Cx43 3′-UTR. Others, such as the DIANA MicroT algorithm, predict the binding of other microRNAs ( 24 ). These si... | 17062625_p21 | 17062625 | DISCUSSION | 4.274966 | biomedical | Study | [
0.9994305968284607,
0.00032618531258776784,
0.00024311061133630574
] | [
0.9994220733642578,
0.00019590885494835675,
0.00030394826899282634,
0.00007815534627297893
] | en | 0.999998 |
Data presented in Figure 3 show that miR-206 expression from pSUPER 206pri is capable of inhibiting the expression of a luciferase reporter bearing the 3′-UTR of Cx43. This experiment demonstrates that even with overexpression of both components of the system, the microRNA is capable of reducing luciferase expression d... | 17062625_p22 | 17062625 | DISCUSSION | 4.246556 | biomedical | Study | [
0.9995427131652832,
0.00025363313034176826,
0.00020367615798022598
] | [
0.9992974996566772,
0.0002496825181879103,
0.00037728447932749987,
0.0000754802895244211
] | en | 0.999994 |
The availability of myoblast cell lines that can recapitulate the myogenic process in vitro has provided a convenient method to study associated developmental processes. In C2C12 cells, myotubes begin to appear at day 2 of differentiation, concurrent with the decline in Cx43 levels. Cx43 protein levels become undetecta... | 17062625_p23 | 17062625 | DISCUSSION | 4.560713 | biomedical | Study | [
0.999219536781311,
0.0005244085332378745,
0.0002561099245212972
] | [
0.9986212253570557,
0.00046350309276022017,
0.0007300368160940707,
0.00018528838700149208
] | en | 0.999999 |
A number of publications have implicated miR-1 in the regulation of skeletal muscle differentiation in both vertebrates and invertebrates as well as during cardiogenesis in vertebrates ( 23 , 29 , 30 ). Our observation that miR-1 downregulates Cx43 during C2C12 differentiation was somewhat surprising because of the pre... | 17062625_p24 | 17062625 | DISCUSSION | 4.1435 | biomedical | Study | [
0.9996993541717529,
0.00014535484660882503,
0.00015531116514466703
] | [
0.9987103939056396,
0.00036357922363094985,
0.0008540931739844382,
0.0000718908486305736
] | en | 0.999996 |
The observation that Cx43 levels decrease during mammalian perinatal skeletal muscle development was first seen decades ago ( 1 , 2 , 9 , 11 , 14 , 31 , 32 ). Data from only one group have suggested that the downregulation may occur at the post-transcriptional level, although this possibility was not pursued in their w... | 17062625_p25 | 17062625 | DISCUSSION | 4.21367 | biomedical | Study | [
0.9995661377906799,
0.00021476343681570143,
0.00021914260287303478
] | [
0.9987767338752747,
0.00016176438657566905,
0.0009923068573698401,
0.0000691871318849735
] | en | 0.999998 |
The miR-206 gene is located between the polycystic kidney and hepatic disease 1 gene and the interleukin-17 gene in mouse (chr 1), rat (chr 9) and human (chr 6). The genomic site where the pre-miR-206 sequence is located contains another microRNA precursor sequence, miR-133b, located a mere ∼3.8 kb downstream, suggesti... | 17062625_p26 | 17062625 | DISCUSSION | 4.542813 | biomedical | Study | [
0.9993922710418701,
0.0003176173777319491,
0.0002901134721469134
] | [
0.9956849813461304,
0.000494431471452117,
0.0036591843236237764,
0.00016143367975018919
] | en | 0.999999 |
Additional recent studies have demonstrated regulatory pathways responsible for the expression of miR-206 during embryonic development ( 35 , 37 ). This work and the work of others have demonstrated an early expression pattern associated with somites in chicken and mouse embryos and with developing muscle in zebrafish ... | 17062625_p27 | 17062625 | DISCUSSION | 4.148142 | biomedical | Study | [
0.9995800852775574,
0.00017437557107768953,
0.00024555603158660233
] | [
0.9779316782951355,
0.0007259193807840347,
0.021198641508817673,
0.00014380928769242018
] | en | 0.999999 |
Real time PCR (RT-PCR) can rapidly, reproducibly and quantitatively determine changes in gene expression ( 1 ). Although microarray analysis can measure large scale gene expression levels simultaneously, its hybridization-related variation often demands validation by other methods. Routinely, RT-PCR is used to verify t... | 17068075_p0 | 17068075 | INTRODUCTION | 4.161116 | biomedical | Study | [
0.9994729161262512,
0.00019992896704934537,
0.00032722949981689453
] | [
0.9297000169754028,
0.008486329577863216,
0.06154545024037361,
0.000268177391262725
] | en | 0.999995 |
Low cost methods for detecting fluorescent dyes which bind to double stranded DNA, such as SYBR Green, are most widely used and suitable for high throughput screening. Since these dyes are not sequence specific, careful consideration should be given to avoid generating extraneous amplicons. One of the obstacles to high... | 17068075_p1 | 17068075 | INTRODUCTION | 4.45428 | biomedical | Study | [
0.9994680285453796,
0.0003471296513453126,
0.0001848791871452704
] | [
0.9981653094291687,
0.0007081531803123653,
0.0009887982159852982,
0.00013772601960226893
] | en | 0.999997 |
Sequences file (refMrna.zip) and intron/exon information tables (refGene.txt.gz) of 23 463 human and 18 737 mouse RefSeq genes (UCSC hg17 and mm6) were downloaded from UCSC genome browser ( ). | 17068075_p2 | 17068075 | Data processing | 3.205402 | biomedical | Study | [
0.9981479644775391,
0.00033664912916719913,
0.0015154926804825664
] | [
0.5135436654090881,
0.48296239972114563,
0.0023166430182754993,
0.0011772643774747849
] | en | 0.999995 |
To assure amplicons free of repetitive elements and sequences of low complexity ( 10 ), we utilized Biowulf, a high-performance Linux cluster at the National Institutes of Health, to mask the repetitive elements using the RepeatMasker application with built-in MaskerAid ( 11 ). | 17068075_p3 | 17068075 | Data processing | 3.834224 | biomedical | Study | [
0.9990816116333008,
0.00018870987696573138,
0.0007297536940313876
] | [
0.978884220123291,
0.020492490381002426,
0.0004160675744060427,
0.00020714954007416964
] | en | 0.999997 |
Primer3 ( 12 ) was used to design primers for each RefSeq entry, with the following parameters: for intronless genes (5.5% of human RefSeq genes and of 12.4% of mouse RefSeq genes), primers were set to be between 17 and 27 bp with 20 bp as optimum, and melting temperature was set to be between 57 and 63°C with 60°C as ... | 17068075_p4 | 17068075 | Data processing | 4.231132 | biomedical | Study | [
0.9994277358055115,
0.00034680511453188956,
0.00022545206593349576
] | [
0.9959713816642761,
0.0034133170265704393,
0.0004653828509617597,
0.00014990277122706175
] | en | 0.999997 |
The BLAST algorithm was used via the NIH biowulf Linux cluster to evaluate all primers against corresponding RefSeq databases. The criteria for possible mis-priming requires that both primers have at least 15 matches in another RefSeq entry (i.e. expectation value, e < 1) ( 13 , 14 ). The BLAST result revealed that 891... | 17068075_p5 | 17068075 | Data processing | 4.223305 | biomedical | Study | [
0.9994575381278992,
0.0003564573999028653,
0.0001859796466305852
] | [
0.9906919598579407,
0.008030429482460022,
0.0009907042840495706,
0.0002868171432055533
] | en | 0.999996 |
qPrimerDepot can be accessed at or by querying the database with a RefSeq ID or a gene name. Batch query service is available upon request if user provides standard gene name or accession number. Flat files and MySQL dump file which have all primer information are also available upon request. | 17068075_p6 | 17068075 | Data processing | 2.053783 | biomedical | Other | [
0.9794386625289917,
0.003232464659959078,
0.01732894778251648
] | [
0.005339497234672308,
0.99365234375,
0.0004454561276361346,
0.000562619767151773
] | en | 0.999998 |
Reverse transcription was applied with Omniscript RT Kit following manufacturer's protocol (Qiagen). A 20 μl RT reaction included 2 μg Universal reference RNA (Stratagen), 1 μM Oligo-dT primer, 2 μl of 10× RT buffer, 0.5 mM each dNTP, 10 U of RNase inhibitor, 4 U of Omniscript Reverse Transcriptase, and DEPC-treated wa... | 17068075_p7 | 17068075 | Experimental validation | 4.022676 | biomedical | Study | [
0.9989219903945923,
0.0006556536536663771,
0.00042241523624397814
] | [
0.7537045478820801,
0.24376791715621948,
0.0015039484715089202,
0.0010235405061393976
] | en | 0.999998 |
Primer sequences were extracted from our database and synthesized by Integrated DNA Technologies (Coralville, IA, USA). Quantitative RT-PCR was carried out in a DNA Engine Opticon-2 Real Time PCR Detection System (MJ Research). In brief, each 20 μl reaction mix comprises 0.3 μM primers (both 5′ and 3′ primers), 1 μl te... | 17068075_p8 | 17068075 | Experimental validation | 4.097751 | biomedical | Study | [
0.9996026158332825,
0.000209486810490489,
0.00018793914932757616
] | [
0.9973810315132141,
0.002161723095923662,
0.0003569548425730318,
0.00010029217082774267
] | en | 0.999997 |
Our database comprises pre-designed primers for 42 133 mouse and human RefSeq genes ( Table 1 ). For most genes three unique sets of primers are provided (96.3% of total). The database provides a simple user interface where the user may enter either the HUGO approved gene symbol or the RefSeq gene identifier . The data... | 17068075_p9 | 17068075 | RESULTS AND DISCUSSION | 3.997545 | biomedical | Study | [
0.9991638660430908,
0.0005168126081116498,
0.00031938706524670124
] | [
0.6752943992614746,
0.319161057472229,
0.004138320684432983,
0.0014061912661418319
] | en | 0.999997 |
To experimentally evaluate the primer quality, 288 genes were arbitrarily selected from a list of genes known to function in the immune response. 288 primer sets were retrieved from the database and synthesized in 96-well plates. Universal human reference RNA was reverse transcribed and used as template in PCR to exami... | 17068075_p10 | 17068075 | RESULTS AND DISCUSSION | 3.996752 | biomedical | Study | [
0.9995673298835754,
0.00018120255845133215,
0.0002513851213734597
] | [
0.9991880059242249,
0.0005297598545439541,
0.00022376146807800978,
0.00005842507744091563
] | en | 0.999995 |
Given the variation of transcript abundance, melting curve analysis followed by gel electrophoresis has been suggested to verify RT-PCR products ( 6 ). Melting curve analysis revealed that 94.1% generate unique product. Visualization by the less sensitive ethidium bromide stain shows that >70% of the primer sets produc... | 17068075_p11 | 17068075 | RESULTS AND DISCUSSION | 4.170248 | biomedical | Study | [
0.9995465874671936,
0.00024520099395886064,
0.00020816567121073604
] | [
0.9992971420288086,
0.00034499677713029087,
0.00029121042462065816,
0.00006658874190179631
] | en | 0.999997 |
The resistance of most qPrimerDepot primer sets to contaminating input genomic DNA is illustrated in Figure 4 . Here the real-time amplification profiles of three intron-bearing genes (VEFG,VEGFB and VEGFC) was compared to that of three non intron-bearing genes (XCR1, SSTR4 and MC1R) after challenge with increasing con... | 17068075_p12 | 17068075 | RESULTS AND DISCUSSION | 4.099805 | biomedical | Study | [
0.9994485974311829,
0.0002788244455587119,
0.00027261467766948044
] | [
0.9995101690292358,
0.00020783120999112725,
0.00022771717340219766,
0.00005427550422609784
] | en | 0.999997 |
Taking advantage of the intron/exon inventory of RefSeq genes and Primer3, a paradigm primer design tool, we designed primers which are contamination resistant for 99% of human and mouse RefSeq genes ( Table 1 ). Since the majority of the primer sets will amplify desired templates under unified annealing temperatures, ... | 17068075_p13 | 17068075 | CONCLUSION | 4.148656 | biomedical | Study | [
0.9996429681777954,
0.00022202912077773362,
0.0001350377278868109
] | [
0.9977682828903198,
0.0015154170105233788,
0.0006061280146241188,
0.00011012324830517173
] | en | 0.999996 |
DNA polymerase δ (pol δ) has a major and essential role in eukaryotic nuclear DNA replication ( 1 ). Pol δ also performs DNA synthesis during homologous recombination and fills DNA gaps during mismatch repair, long patch base excision repair of damaged bases and nucleotide excision repair of bulky DNA lesions [reviewed... | 16936322_p0 | 16936322 | INTRODUCTION | 4.354514 | biomedical | Study | [
0.9992204904556274,
0.00038552944897674024,
0.0003938662412110716
] | [
0.7913095355033875,
0.0015327547444030643,
0.20670540630817413,
0.0004523451207205653
] | en | 0.999997 |
In performing its roles in replication and repair, pol δ is assisted by accessory proteins. The three-subunit replication protein A (RPA) complex ( 6 ) binds single-stranded DNA and coordinates the exchange of pol δ and other proteins at template–primer termini ( 7 ). In addition, the processivity of pol δ is enhanced ... | 16936322_p1 | 16936322 | INTRODUCTION | 4.660524 | biomedical | Study | [
0.997572124004364,
0.0012702889507636428,
0.0011576216202229261
] | [
0.5802931785583496,
0.0019176877103745937,
0.41673606634140015,
0.0010530196595937014
] | en | 0.999998 |
Here we test these ideas by examining the fidelity of DNA synthesis by three-subunit yeast DNA polymerase δ alone and its fidelity in the presence of RPA alone, PCNA (plus its loader RFC) and all three accessory protein complexes. To evaluate the effects of these accessory proteins on both nucleotide selectivity and pr... | 16936322_p2 | 16936322 | INTRODUCTION | 4.33088 | biomedical | Study | [
0.9992882609367371,
0.0004586867871694267,
0.0002530812635086477
] | [
0.9983721375465393,
0.000262034242041409,
0.0012530818348750472,
0.00011279070895398036
] | en | 0.999998 |
Yeast RPA, PCNA and RFC were purified from Escherichia coli overproducing strains as described elsewhere ( 24 , 25 ). All materials for the fidelity assay were from previously described sources ( 23 , 26 ). | 16936322_p3 | 16936322 | Materials | 3.551251 | biomedical | Study | [
0.9984921216964722,
0.00023108458844944835,
0.001276839291676879
] | [
0.9798737168312073,
0.01843363791704178,
0.0014449912123382092,
0.0002477301750332117
] | en | 0.999995 |
Plasmid pBL335 (bluescript, 2 µM ori, TRP1 , M13 ori, GAL1-10 GST-POL3 ) contains the Schistosoma japanicum glutathione S -transferase gene (GST) fused to the N-terminus of the POL3 gene in vector pRS424-GALGSTPKA. The GST tag is separated from the POL3 gene by a recognition sequence for the human rhinoviral protease (... | 16936322_p4 | 16936322 | Overexpression and purification of Pol δ | 4.345788 | biomedical | Study | [
0.9990509152412415,
0.0005788258276879787,
0.0003702749090734869
] | [
0.9963139891624451,
0.003142179688438773,
0.00031859276350587606,
0.00022535726020578295
] | en | 0.999996 |
Overexpression was in Saccharomyces cerevisiae strain BJ2168 transformed with pBL341 and with either pBL335 or pBL335-DV. Growth and induction was as described elsewhere, and so was the preparation of cell lysates by blending with dry ice ( 29 ). Cells (60 g of packed cells resuspended in 20 ml of water) frozen previou... | 16936322_p5 | 16936322 | Overexpression and purification of Pol δ | 4.37264 | biomedical | Study | [
0.9989776611328125,
0.0006788953905925155,
0.0003434818936511874
] | [
0.9978042244911194,
0.0015380593249574304,
0.0004629317845683545,
0.0001948211865965277
] | en | 0.999996 |
Reactions (25 µl) contained 20 mM Tris–HCl (pH 7.7), 8 mM MgAc 2 , 75 mM NaCl, 0.5 mM ATP, 100 µM of each dNTP, 1 mM DTT, 100 mg/ml BSA and 40 fmol (1.6 nM) gapped M13mp2 DNA. When included, the amounts of the accessory proteins used were 500 fmol PCNA, 200 fmol RFC and 10 pmol RPA, an amount more than sufficient to co... | 16936322_p6 | 16936322 | Gap-filling DNA synthesis reactions and product analysis | 4.226756 | biomedical | Study | [
0.999423623085022,
0.0003474555560387671,
0.0002289632539032027
] | [
0.9992350339889526,
0.0003723388654179871,
0.00030839364626444876,
0.00008420141239184886
] | en | 0.999998 |
DNA products of gap-filling reactions were introduced into E.coli cells and plated as described elsewhere ( 23 ) to score blue M13 plaques (correct synthesis) and light blue and colorless plaques (containing errors). The types of errors were determined by sequencing the lacZ α-complementation gene in single-stranded DN... | 16936322_p7 | 16936322 | Gap-filling DNA synthesis reactions and product analysis | 4.108135 | biomedical | Study | [
0.9994716048240662,
0.0002644987544044852,
0.00026389784761704504
] | [
0.9993495345115662,
0.0002585139009170234,
0.0003370384802110493,
0.000054967393225524575
] | en | 0.999996 |
Pol δ fidelity with and without accessory proteins was determined for synthesis to fill a single-stranded gap in a circular duplex M13mp2 DNA substrate. This gap contains the lacZ α-complementation template sequence that when copied correctly results in a blue M13 plaque phenotype. Polymerization errors are detected as... | 16936322_p8 | 16936322 | Fidelity measurements and calculation of error rates | 4.246671 | biomedical | Study | [
0.9993785619735718,
0.0003755320212803781,
0.00024588964879512787
] | [
0.9993340373039246,
0.0002266761293867603,
0.00035289369407109916,
0.00008638112194603309
] | en | 0.999997 |
In order to separate the effects of the accessory factors on polymerase insertion fidelity from those on proofreading efficiency, we will first discuss our results with the exonuclease-deficient pol δ, followed by those with the wild-type enzyme. Because similar results were obtained for each of the two exonuclease-def... | 16936322_p9 | 16936322 | Fidelity measurements and calculation of error rates | 3.954427 | biomedical | Study | [
0.9994125366210938,
0.0002293591242050752,
0.00035815907176584005
] | [
0.9984173774719238,
0.0007181827677413821,
0.0007872851565480232,
0.00007708404882578179
] | en | 0.999996 |
The most common errors generated by pol δ were single base substitutions. The calculated average single base substitution error rate of exonuclease-deficient pol δ alone is 6.3 × 10 −5 ( Table 2 ), confirming an earlier report that yeast pol δ has high base substitution fidelity ( 5 ). Interestingly, similar base subst... | 16936322_p10 | 16936322 | Effects on selectivity against base–base mismatches | 4.197137 | biomedical | Study | [
0.99945467710495,
0.00030549027724191546,
0.00023984794097486883
] | [
0.9993897676467896,
0.0002459139795973897,
0.00029660225845873356,
0.00006767040758859366
] | en | 0.999997 |
The base substitution values in Table 2 are average error rates for numerous different mismatches in a variety of sequence contexts. From these average rates, it cannot be excluded that the accessory proteins have modest effects on nucleotide selectivity and/or mismatch extension for specific mismatches and/or in speci... | 16936322_p11 | 16936322 | Effects on selectivity against base–base mismatches | 4.304262 | biomedical | Study | [
0.9994376301765442,
0.0003007578488904983,
0.000261655542999506
] | [
0.99919193983078,
0.00027821792173199356,
0.0004539591900538653,
0.00007588072185171768
] | en | 0.999997 |
An estimate of the contribution of proofreading to base substitution fidelity in the absence and presence of the accessory proteins can be obtained by comparing error rates for exonuclease-deficient pol δ to those for wild-type pol δ ( Table 2 ). We believe that these are minimal estimates because the base substitution... | 16936322_p12 | 16936322 | Effects on proofreading of base–base mismatches | 4.30914 | biomedical | Study | [
0.9993759989738464,
0.0003786291053984314,
0.0002454398781992495
] | [
0.9992375373840332,
0.00026505341520532966,
0.00040180422365665436,
0.00009564169158693403
] | en | 0.999997 |
The second most common single base error made by pol δ is deletion of 1 nt ( Table 2 ). The rates at which these errors are generated by pol δ in the absence or presence of RFC/PCNA and/or RPA either do not differ in a statistically significant manner, or they differ in a statistically significant manner but by <2-fold... | 16936322_p13 | 16936322 | Effects on single nucleotide deletion and addition errors | 4.228887 | biomedical | Study | [
0.999363124370575,
0.00035503454273566604,
0.0002819089568220079
] | [
0.9993101358413696,
0.0003247549175284803,
0.0002893579949159175,
0.00007569317676825449
] | en | 0.999998 |
Exonuclease-deficient pol δ also generates single base additions at a readily detectable overall average rate of 0.66 × 10 −5 ( Table 2 ). This rate is not appreciably influenced by RPA (0.80 × 10 −5 ), but is reduced ∼7-fold by PCNA plus RFC . Wild-type pol δ alone (error rate ≥0.026 × 10 −5 ) is at least 25-fold more... | 16936322_p14 | 16936322 | Effects on single nucleotide deletion and addition errors | 4.260118 | biomedical | Study | [
0.9993782043457031,
0.0003368646721355617,
0.0002848147414624691
] | [
0.9993047714233398,
0.0003400136192794889,
0.000274633668595925,
0.00008061242260737345
] | en | 0.999997 |
The effects of accessory proteins on single base error rates are modest in comparison with the dominant role of the polymerase itself in discriminating against single base errors. However, the situation is different for deletions of larger numbers of nucleotides located between direct repeat sequences. We found previou... | 16936322_p15 | 16936322 | Effects on deletions between direct repeats | 4.227528 | biomedical | Study | [
0.9994602799415588,
0.0003055566630791873,
0.0002341776853427291
] | [
0.9993498921394348,
0.00020924358977936208,
0.000367062195437029,
0.00007378601731033996
] | en | 0.999998 |
How RPA and PCNA may suppress formation of large deletions by pol δ can be considered in light of a model for deletions between direct repeats [ Figure 2 ( 39 )]. In this model, after the first repeat sequence encountered by the polymerase is copied, the primer frays and then relocates to the second repeat sequence. Hy... | 16936322_p16 | 16936322 | Effects on deletions between direct repeats | 4.737188 | biomedical | Study | [
0.9987497329711914,
0.0008398336940445006,
0.0004103759420104325
] | [
0.9962372779846191,
0.0010342964669689536,
0.002378112869337201,
0.00035032755113206804
] | en | 0.999997 |
The accuracy of intron excision from pre-mRNAs relies on the precise recognition of rather degenerate splicing signals. Splice sites are defined by consensus sequences where only the two terminal nucleotides [GT at the acceptor (5′) and AG at the donor (3′) splice sites for the majority of introns] are highly conserved... | 16936312_p0 | 16936312 | INTRODUCTION | 4.917306 | biomedical | Study | [
0.9972122311592102,
0.001499249367043376,
0.0012885017786175013
] | [
0.7512384057044983,
0.003608601400628686,
0.24382120370864868,
0.0013318239944055676
] | en | 0.999996 |
Alternative splicing is an important mechanism for the generation of proteome complexity and fine tuning of gene expression at the post-transcriptional level both in the metazoan and plant species. Differential intron removal allows production of splice variants which may code for distinct protein isoforms affecting th... | 16936312_p1 | 16936312 | INTRODUCTION | 4.274549 | biomedical | Study | [
0.9984182119369507,
0.0007500601350329816,
0.0008317132014781237
] | [
0.452223002910614,
0.09580313414335251,
0.4504108130931854,
0.0015631492715328932
] | en | 0.999996 |
Our analysis of the Arabidopsis genome has revealed 19 SR proteins which is almost twice as much as in humans ( 5 , 6 ). They fall into seven subfamilies ( 6 ) some of which have orthologues in metazoan (SF2/ASF, 9G8, SC35) but interestingly three of them seem to be plant-specific (RS, RS2Z and SCL). Many of the SR pro... | 16936312_p2 | 16936312 | INTRODUCTION | 4.487518 | biomedical | Study | [
0.9991469383239746,
0.0004139212251175195,
0.00043918672599829733
] | [
0.9989853501319885,
0.0003690442827064544,
0.0005295180599205196,
0.0001161774416686967
] | en | 0.999996 |
In this study, we extend the analysis of alternative splicing and its regulation to the plant-specific RS subfamily, focusing on atRSp31 . We also show that atRSZ33 , a member of the RS2Z subfamily, is involved in this regulation. In addition, we show that the position of the long intron and its capacity for alternativ... | 16936312_p3 | 16936312 | INTRODUCTION | 4.204835 | biomedical | Study | [
0.9991374015808105,
0.00033018813701346517,
0.000532439153175801
] | [
0.9995300769805908,
0.00017079136159736663,
0.00024647609097883105,
0.000052569354011211544
] | en | 0.999997 |
Arabidopsis and rice sequences can be found at and under following numbers: atRSp31 , atRSp31a , atRSp40 , atRSp41 , osRSp29 , osRSp33 , atRSZ32 , atRSZ33 , osRSZ36 , osRSZ37a , osRSZ37b and osRSZ39 . Accession numbers of Arabidopsis , rice, maize, Pinus taeda , Physcomitrella patens and Chlamydomonas reinhardtii trans... | 16936312_p4 | 16936312 | Accession numbers | 1.761809 | biomedical | Other | [
0.9825687408447266,
0.0009179117623716593,
0.01651330105960369
] | [
0.09299678355455399,
0.9045491814613342,
0.0013974595349282026,
0.0010566443670541048
] | en | 0.999997 |
To identify splice variants of Arabidopsis SR proteins, corresponding genomic sequences were used in BLAST search against EST database limited to Arabidopsis thaliana at . ESTs with matches to corresponding gene were selected and then aligned using Geneseqer ( 18 ) at . | 16936312_p5 | 16936312 | Sequence retrieval and analysis | 3.892373 | biomedical | Study | [
0.9991328120231628,
0.00017631259106565267,
0.0006909661460667849
] | [
0.9978062510490417,
0.0018866690807044506,
0.0002415578783256933,
0.00006550041143782437
] | en | 0.999996 |