Abstract:
:A single-stage polymerase-based procedure is described that allows extensive modifications of DNA. The version described here uses the QuikChange Site-Directed Mutagenesis System kit supplied by Stratagene. The original protocol is replaced by a single-stage method in which linear production of complementary strands is accomplished in separate single primer reactions. This has proved effective in introducing insertions and deletions into large gene/vector combinations without subcloning.
journal_name
Mol Biotechnoljournal_title
Molecular biotechnologyauthors
Salerno JC,Jones RJ,Erdogan E,Smith SMdoi
10.1385/MB:29:3:225keywords:
subject
Has Abstractpub_date
2005-03-01 00:00:00pages
225-32issue
3eissn
1073-6085issn
1559-0305pii
MB:29:3:225journal_volume
29pub_type
杂志文章abstract::IgG class antibodies express catalytic activities rarely and at very low levels. Here, we studied polyclonal IgA and IgG preparations from healthy human sera and saliva for the ability to hydrolyze model peptidyl-aminomethylcoumarin (peptide-AMC) substrates. These substrates permit objective evaluation of the catalyti...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-007-0003-7
更新日期:2007-06-01 00:00:00
abstract::Monoclonal antibodies (mAbs) are the fastest growing class of biopharmaceuticals reflecting their diverse applications in research and the clinic. The correct glycosylation of mAbs is required to elicit effector functions such as complement-dependent and antibody-dependent cell-mediated cytotoxicity, although these ma...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-012-9531-x
更新日期:2013-03-01 00:00:00
abstract::ATP-binding cassette transporter G2 (ABCG2) gene encodes a protein that has a wide variety of substrates and is responsible for the active secretion of clinically and toxicologically important molecules into milk. Although known in many species, this marks the first time this gene product has been reported in goats. I...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-007-9024-5
更新日期:2008-05-01 00:00:00
abstract::Artificial microRNA (amiRNA) has become a powerful tool for gene silencing in plants. A new method for easy and rapid construction of rice artificial miRNA vector is described. The procedure involved modification of the pCAMBIA1300-UR vector by insertion of a 'vector modification fragment'. This was prepared from the ...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-010-9291-4
更新日期:2010-11-01 00:00:00
abstract::In the present study, four full-length Dof (DNA-binding with one finger) genes from Sorghum bicolor namely SbDof1, SbDof19, SbDof23, and SbDof24 were PCR amplified, gel eluted, cloned, and sequenced (accession number HQ540084, HQ540085, HQ540086, and HQ540087, respectively). These sequences were further characterized ...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-012-9536-5
更新日期:2013-05-01 00:00:00
abstract::UV-B radiation can damage biomolecules, such as DNA, RNA, and proteins, halting essential cellular processes; this damage is partly due to ROS generation. Plant secondary metabolites may protect against UV-B. Psychotria brachyceras Müll. Arg. (Rubiaceae), a subtropical shrub, produces brachycerine, a monoterpene indol...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-012-9546-3
更新日期:2013-05-01 00:00:00
abstract::This review outlines the various methods available for submitting sequence data to the EMBL Nucleotide Sequence Database. Depending on the type of sequence data and the facilities available to the submitter, one method may be more suitable than another. Recent developments have been the World Wide Web submission tool,...
journal_title:Molecular biotechnology
pub_type: 杂志文章,评审
doi:10.1007/BF02762322
更新日期:1996-08-01 00:00:00
abstract::In this study, we investigated the relative expression of the Rous sarcoma virus (RSV) promoter-driven expression of enhanced green fluorescent protein (EGFP) in fibroblasts of transgenic quails. We analyzed the direct influence of CpG methylation of the RSV promoter on the transcriptional activity of delivered transg...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-011-9393-7
更新日期:2011-11-01 00:00:00
abstract::D-Allulose is a rare monosaccharide that exists in extremely small quantities in nature, and it is also hard to prepare at a large scale via chemical or enzyme synthetic route due to low conversion and downstream separation complexity. Using D-psicose epimerase and L-rhamnulose kinase, a method enabling high conversio...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-019-00174-6
更新日期:2019-06-01 00:00:00
abstract::In order to further study the B subunit of the Escherichia coli heat-labile enterotoxin (LTB), we obtained the LTB gene from pathogenic E. coli, cloned it into the pET22b (+) prokaryotic expression vector, and expressed it as a fusion protein with His tag in E. coli BL21 (DE3). The recombinant LTB was expressed and pu...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-008-9101-4
更新日期:2009-02-01 00:00:00
abstract::In the context of clinical gene transfer using viral vectors, the risk of memory antivector immunity is often poorly appreciated. The immunological past of the patient, the site of injection, and the vector dose will play intertwined and decisive roles in the safety and efficacy of treatment. To circumvent the drawbac...
journal_title:Molecular biotechnology
pub_type: 杂志文章,评审
doi:10.1385/MB:34:2:247
更新日期:2006-10-01 00:00:00
abstract::Brn-3a, a member of the POU gene family (so-called because of the similarity with the group of transcription factors Pit, Oct, and Unc), was found in neuronal cells engaged in the transcription activity of the p1 and p2 promoters of the most powerful antiapoptotic gene, namely, Bcl-2. The alternative splicing of Brn-3...
journal_title:Molecular biotechnology
pub_type: 杂志文章,评审
doi:10.1385/MB:22:2:123
更新日期:2002-10-01 00:00:00
abstract::We report a rapid and simple method for isolating the 5'-end of plant genes from genomic DNA by polymerase chain reaction (PCR) with TATA-box-based degenerate primers (TDPs). The TDPs were specially designed according to the TATA box, which is conserved in the promoter region of most plant genes. The unknown 5'-ends o...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-010-9323-0
更新日期:2011-02-01 00:00:00
abstract::PREP1/p160 is a protein complex with relevant physiopathological roles in vivo. p160 regulates PREP1 transcriptional activity by preventing the formation of other PREP1-containing complexes, whereas PREP1 regulates p160 activity by increasing its stability. This induces the repression of the insulin-regulated glucose ...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-016-9932-3
更新日期:2016-05-01 00:00:00
abstract::The MYB proteins constitute one of the largest transcription factor families in plants. Much research has been performed to determine their structures, functions, and evolution, especially in the model plants, Arabidopsis, and rice. However, this transcription factor family has been much less studied in wheat (Triticu...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-011-9486-3
更新日期:2012-10-01 00:00:00
abstract::Coptotermes curvignathus is a termite that, owing to its ability to digest living trees, serves as a gold mine for robust industrial enzymes. This unique characteristic reflects the presence of very efficient hydrolytic enzyme systems including cellulases. Transcriptomic analyses of the gut of C. curvignathus revealed...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-017-0015-x
更新日期:2017-07-01 00:00:00
abstract::We investigated the efficiency of a single plasmid transfection along the longitudinal axis of the regenerating soleus of young rats. This also reflected transfection efficiency along the fibers because the soleus is a nearly fusiform muscle in young animals. The complete regeneration was induced by notexin and the tr...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-012-9555-2
更新日期:2013-06-01 00:00:00
abstract::Agrobacterium-mediated transformation is the method of choice to engineer desirable genes into plants. Here we describe a protocol for demonstrating T-DNA transfer from Agrobacterium into the economically important graminaceous plant maize. Expression of the T-DNA-located GUS gene was observed with high efficiency on ...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1385/MB:13:2:165
更新日期:1999-12-01 00:00:00
abstract::Since it was first introduced in late 1990s Wave bioreactor has been used for protein production by mammalian and insect cell lines. However, using Wave bioreactor to produce human monoclonal antibody by stable Drosophila Schneider 2 (S2) cell transfectants has not been reported before. In this study, S2 cells were co...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-011-9484-5
更新日期:2012-10-01 00:00:00
abstract::Association studies using genome scans to identify quantitative trait loci for multifactorial disorders, with anything approaching reasonable power, have been compromised by the need for a very dense array of genetic markers and large numbers of affected individuals. These requirements impose enormous burdens on the g...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1385/mb:22:3:253
更新日期:2002-11-01 00:00:00
abstract::The simple use of nonisotopic hybridization probes to detect complementary sequences provides valuable information in a large number of research and commercial applications. In hybridization assays, the four "S's (speed, simplicity, sensitivity, and specificity) are important criteria for determining the choice of pro...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/BF02740771
更新日期:1996-10-01 00:00:00
abstract::High-quality RNA preparations are critical for further applications such as reverse transcriptase-polymerase chain reaction (RT-PCR) transcript amplifications, and elaboration of cDNA and expressed sequence tag libraries. Melanins are phenolic compounds present in many fungi and apparently play key roles in fungi path...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1385/MB:34:1:45
更新日期:2006-09-01 00:00:00
abstract::The extracellular signal-regulated kinase (ERK) cascade is a central intracellular signaling pathway that is activated by a variety of extracellular stimuli, and thereby regulates cellular processes such as proliferation, differentiation, and oncogenic transformation. To execute these functions, the signals of those s...
journal_title:Molecular biotechnology
pub_type: 杂志文章,评审
doi:10.1385/MB:29:1:57
更新日期:2005-01-01 00:00:00
abstract::Adenoviral dodecahedron (Dd) is a virus-like particle composed of twelve pentameric penton base (Pb) proteins, responsible for adenovirus cell penetration. It is generated spontaneously in the baculovirus system upon expression of the Pb gene of adenovirus serotype 3. This particle shows remarkable cell penetration ab...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-015-9850-9
更新日期:2015-06-01 00:00:00
abstract::Enamel is the covering tissue of teeth, made of regularly arranged hydroxyapatite crystals deposited on an organic matrix composed of 90% amelogenin that is completely degraded at the end of the enamel formation process. Amelogenin has a biomineralizing activity, forming nanoparticles or nanoribbons that guide hydroxy...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-017-0030-y
更新日期:2017-10-01 00:00:00
abstract::This article describes the construction, expression, and purification of RNase single-chain antibody fusion proteins. To construct a fusion protein, the gene for each moiety, the RNase and the binding ligand, is modified separately to contain complementary DNA encoding a 13 amino acid spacer that separates the RNase f...
journal_title:Molecular biotechnology
pub_type: 杂志文章,评审
doi:10.1385/MB:20:1:063
更新日期:2002-01-01 00:00:00
abstract::An endochitinase gene from the Serratia marcescens Nima strain (chiA Nima) was cloned, sequenced, and expressed in Escherichia coli DH5alphaF', and the recombinant protein (ChiA Nima) was purified by hydrophobic interaction chromatography. chiA Nima contains an open reading frame (ORF) that encodes an endochitinase wi...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1385/MB:31:2:103
更新日期:2005-10-01 00:00:00
abstract::An alternative method to combine mutagenesis PCR with dITP and fragmentation by endonuclease V for directed evolution was developed. In comparison to the routine protocol for directed evolution, dITP was used as mutation reagent in the mutagenesis PCR. Subsequently, the incorporated dITP in the PCR products could repr...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-012-9516-9
更新日期:2013-01-01 00:00:00
abstract::The diversity and complexity of bioinformatics tools currently available for protein sequence analysis can make it difficult to know where to begin when presented with a new sequence. In this article, we present a protocol outlining one approach to sequence analysis that should give as comprehensive a picture as possi...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1007/s12033-007-0032-2
更新日期:2007-07-01 00:00:00
abstract::A sensitive method for staining proteins after transfer from polyacrylamide gels to nitrocellulose paper is described. Transferred proteins are first derivatized by reaction of the nitrocellulose replica with sulfosuccinimidobiotin and are then reacted sequentially with streptavidin, rabbit anti-streptavidin, and hors...
journal_title:Molecular biotechnology
pub_type: 杂志文章
doi:10.1385/MB:15:2:161
更新日期:2000-06-01 00:00:00