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Shilpa Nagaraju, Naomi Kathleen Davies, David Jeffrey Fraser Walker, Michael Köpke, Séan Dennis Simpson
BACKGROUND: Impactful greenhouse gas emissions abatement can now be achieved through gas fermentation using acetogenic microbes for the production of low-carbon fuels and chemicals. However, compared to traditional hosts like Escherichia coli or yeast, only basic genetic tools exist for gas-fermenting acetogens. To advance the process, a robust genetic engineering platform for acetogens is essential. RESULTS: In this study, we report scarless genome editing of an industrially used model acetogen, Clostridium autoethanogenum, using the CRISPR/Cas9 system...
2016: Biotechnology for Biofuels
Ruchi Goel, Smriti Nagpal, Sushil Kumar, Ravi Meher, Saurabh Kamal, Sonam Garg
PURPOSE: To evaluate and compare the success rate of transcanalicular laser-assisted dacryocystorhinostomy with endonasal augmentation, with and without intubation, in patients suffering from primary acquired nasolacrimal duct obstruction, at 1 year of follow up. METHODS: A prospective, randomized interventional pilot study was conducted at a tertiary care center, in accordance with the guidelines of Declaration of Helsinki. Sixty eyes of 60 adult patients with primary acquired nasolacrimal duct obstruction were included...
October 20, 2016: Ophthalmic Plastic and Reconstructive Surgery
Zhongqiu Chen, Wen Ling, Guangdong Shang
Pseudomonas putida KT2440 is a saprophytic, generally recognized as safe microorganism that plays important roles in the biodegradation and production of value-added chemicals. Chromosomal gene deletion of P. putida KT2440 usually involves time-consuming gene coning, conjugal transfer and counterselection. Recently, we developed a P. putida KT2440 markerless gene deletion method based on recombineering and Cre/loxP site-specific recombination. PCR-based λ Red recombineering circumvents the tedious cloning steps and is more amenable to high-throughput manipulation...
October 7, 2016: FEMS Microbiology Letters
Dirk Landgraf, Dann Huh, Erinc Hallacli, Susan Lindquist
Gene tagging with fluorescent proteins is commonly applied to investigate the localization and dynamics of proteins in their cellular environment. Ideally, a fluorescent tag is genetically inserted at the endogenous locus at the N- or C- terminus of the gene of interest without disrupting regulatory sequences including the 5' and 3' untranslated region (UTR) and without introducing any extraneous unwanted "scar" sequences, which may create unpredictable transcriptional or translational effects. We present a reliable, low-cost, and highly efficient method for the construction of such scarless C-terminal and N-terminal fusions with fluorescent proteins in yeast...
2016: PloS One
Annika Ciragan, A Sesilja Aranko, Igor Tascon, Hideo Iwaï
Intervening protein sequences (inteins) from extremely halophilic haloarchaea can be inactive under low salinity but could be activated by increasing salt contents to a specific concentration for each intein. The halo-obligatory inteins confer high solubility under both low and high salinity conditions. We showed the broad utility of salt-dependent protein splicing in cis and trans by demonstrating backbone cyclization, self-cleavage for purification, and scarless protein ligation for segmental isotopic labeling...
October 6, 2016: Journal of Molecular Biology
Fufang Ni, Sai Luo, Dongmei Yu, Yangdong Zhu, Yong Shang, Yida Chen, Haiqian Xu, Qing Wang, Lijun Hao
BACKGROUND: The presence of mild or moderate medial epicanthus is typical in Asian patients. Numerous epicanthoplasty techniques have been described previously. However, these methods usually leave obvious scars in the medial canthal area. The aim of this report is to introduce a novel epicanthoplasty technique and a concomitant double eyelidplasty which avoid leaving scars in the medial canthal region. METHODS: From July 2013 to July 2015, 252 patients received epicanthoplasty and concomitant double eyelidplasty with this new technique...
October 4, 2016: Aesthetic Plastic Surgery
Omar E Aly, Douglas H Black, Haroon Rehman, Irfan Ahmed
BACKGROUND: Appendicectomy is a well-established surgical procedure used in the management of acute appendicitis. The operation can be performed with minimally invasive surgery or as an open procedure. A further development in the minimally invasive appendicectomy technique has been the introduction of single incision laparoscopic surgery (SILA). AIM: To ascertain any differences in outcomes from available trials comparing SILA with conventional multi-incision laparoscopic appendicectomy (CLA)...
October 2, 2016: International Journal of Surgery
Zhong Zheng, Xinli Zhang, Catherine Dang, Steven Beanes, Grace X Chang, Yao Chen, Chen-Shuang Li, Kevin S Lee, Kang Ting, Chia Soo
In contrast to adult and late-gestation fetal skin wounds, which heal with scar, early-gestation fetal skin wounds display a remarkable capacity to heal scarlessly. Although the underlying mechanism of this transition from fetal-type scarless healing to adult-type healing with scar has been actively investigated for decades, in utero restoration of scarless healing in late-gestation fetal wounds has not been reported. In this study, using loss- and gain-of-function rodent fetal wound models, we identified that fibromodulin (Fm) is essential for fetal-type scarless wound healing...
September 22, 2016: American Journal of Pathology
A de Runz, D Boccara, F Bekara, M Chaouat, M Mimoun
BACKGROUND: Delayed breast reconstruction with tissue expansion may be risky after radiotherapy, due to the poor skin quality. To permit the use of the tissue expansion procedure, we propose a scarless latissimus dorsi flap to bring tissue trophicity, by a healthy vascularized muscular interface with no donor scar and no patch effect. The objective of this study is to assess the outcome of the tissue expansion technique with scarless latissimus dorsi flap after post-mastectomy radiotherapy...
September 7, 2016: Annales de Chirurgie Plastique et Esthétique
Cecilia W P Li-Tsang
No abstract text is available yet for this article.
2016: Burns and trauma
Ran Yu, Yumei Ding, Lijuan Zhu, Yinying Qu, Chenguang Zhang, Lin Liu, Lili Chen
OBJECTIVE: Wounds are common in the oral cavity. During wound healing, several cytokines are released, which are probably helpful in providing wound debridement, removal of damaged tissues and microbes. Most of the target cells of IL-22 are epithelial cells, which play an important role in mucosa immunity. DESIGN: The function of IL-22 in oral diseases is not well understood. We investigated the expression level of IL-22, collagen I and p-stat3 (Tyr705) via a mice tongue wound model in vivo and detected the effect of IL-22 on the expression of MMP-1, type I collagen and p-stat3 in keratinocytes...
August 2, 2016: Archives of Oral Biology
Brian Chaikind, Jeffrey L Bessen, David B Thompson, Johnny H Hu, David R Liu
We describe the development of 'recCas9', an RNA-programmed small serine recombinase that functions in mammalian cells. We fused a catalytically inactive dCas9 to the catalytic domain of Gin recombinase using an optimized fusion architecture. The resulting recCas9 system recombines DNA sites containing a minimal recombinase core site flanked by guide RNA-specified sequences. We show that these recombinases can operate on DNA sites in mammalian cells identical to genomic loci naturally found in the human genome in a manner that is dependent on the guide RNA sequences...
August 11, 2016: Nucleic Acids Research
Rosana Mateu, Veronika Živicová, Eliška Drobná Krejčí, Miloš Grim, Hynek Strnad, Čestmír Vlček, Michal Kolář, Lukáš Lacina, Peter Gál, Jiří Borský, Karel Smetana, Barbora Dvořánková
Clinical evidence suggests that healing is faster and almost scarless at an early neonatal age in comparison with that in adults. In this study, the phenotypes of neonatal and adult dermal fibroblasts and keratinocytes (nestin, smooth muscle actin, keratin types 8, 14 and 19, and fibronectin) were compared. Furthermore, functional assays (proliferation, migration, scratch wound closure) including mutual epithelial‑mesenchymal interactions were also performed to complete the series of experiments. Positivity for nestin and α smooth muscle actin was higher in neonatal fibroblasts (NFs) when compared with their adult counterparts (adult fibroblasts; AFs)...
October 2016: International Journal of Molecular Medicine
Justyna Podolak-Popinigis, Anna Ronowicz, Monika Dmochowska, Agnieszka Jakubiak, Paweł Sachadyn
AIM: Fetal skin is known to heal without scarring. In mice, the phenomenon is observed until the 16-17 day of gestation - the day of transition from scarless to normal healing. The study aims to identify key methylome and transcriptome changes following the transition. MATERIALS & METHODS: Methylome and transcriptome profiles were analyzed in murine dorsal skin using microarray approach. RESULTS & CONCLUSION: The genes associated with inflammatory response and hyaluronate degradation showed increased DNA methylation before the transition, while those involved in embryonic morphogenesis, neuron differentiation and synapse functions did so after...
October 2016: Epigenomics
Yu Tian, Shuodong Wu, Chun-Chih Chen, Yongsheng Chen
BACKGROUND: Optimization of quality of life is an important goal in the advanced surgery. Herein we share our fifteen cases to demonstrate the feasibility of scarless laparoendoscopic single-site common bile duct (CBD) exploration using conventional instruments in the management of CBD stones. METHODS: Fifteen patients (4 male) underwent transumbilical laparoendoscopic single-site cholecystectomy and CBD exploration: the patients were diagnosed with CBD stones. The endoscopic nasobiliary drainage (ENBD), endoscopic retrograde biliary drainage (ERBD) or J-type tube was introduced into the CBD preoperatively or intraoperatively...
September 2016: International Journal of Surgery
Abigail M Lamb, Elizabeth A Walker, Patricia J Wittkopp
Genome editing via the CRISPR/Cas9 RNA-guided nuclease system has opened up exciting possibilities for genetic analysis. However, technical challenges associated with homology-directed repair have proven to be roadblocks for producing changes in the absence of unwanted, secondary mutations commonly known as "scars". To address these issues, we developed a two-stage, marker-assisted strategy to facilitate precise, "scarless" edits in Drosophila with a minimal requirement for molecular screening. Using this method, we modified two base pairs in a gene of interest without altering the final sequence of the CRISPR cut sites...
August 5, 2016: Fly
Brian R Shy, Matthew S MacDougall, Ryan Clarke, Bradley J Merrill
CRISPR/Cas9 nucleases have enabled powerful, new genome editing capabilities; however, the preponderance of non-homologous end joining (NHEJ) mediated repair events over homology directed repair (HDR) in most cell types limits the ability to engineer precise changes in mammalian genomes. Here, we increase the efficiency of isolating precise HDR-mediated events in mouse embryonic stem (ES) cells by more than 20-fold through the use of co-incidental insertion (COIN) of independent donor DNA sequences. Analysis of on:off-target frequencies at the Lef1 gene revealed that bi-allelic insertion of a PGK-Neo cassette occurred more frequently than expected...
September 19, 2016: Nucleic Acids Research
Toshio Sawai, Takeo Yonekura, Katsuji Yamauchi, Takuya Kimura, Keisuke Nose
We present a new, scarless, circular incisional approach around the umbilical cord for neonates with intestinal atresia. This novel approach achieves truly woundless surgery. It is simple, safe, and can be used for an intestinal surgical treatment in neonates.
October 2016: Pediatric Surgery International
Mu-En Chung, I-Hsin Yeh, Li-Yu Sung, Meng-Ying Wu, Yun-Peng Chao, I-Son Ng, Yu-Chen Hu
Metabolic engineering often necessitates chromosomal integration of multiple genes but integration of large genes into Escherichia coli remains difficult. CRISPR/Cas9 is an RNA-guided system which enables site-specific induction of double strand break (DSB) and programmable genome editing. Here, we hypothesized that CRISPR/Cas9-triggered DSB could enhance homologous recombination and augment integration of large DNA into E. coli chromosome. We demonstrated that CRISPR/Cas9 system was able to trigger DSB in >98% of cells, leading to subsequent cell death, and identified that mutagenic SOS response played roles in the cell survival...
July 25, 2016: Biotechnology and Bioengineering
Ting Gong, Ruihua Liu, You Che, Xiaoqing Xu, Fengjie Zhao, Huilei Yu, Cunjiang Song, Yanping Liu, Chao Yang
Currently, chlorpyrifos (CP) and carbofuran are often applied together to control major agricultural pests in many developing countries, in most cases, they are simultaneously detected in agricultural soils. Some cost-effective techniques are required for the remediation of combined pollution caused by multiple pesticides. In this work, we aim at constructing a detectable recombinant microorganism with the capacity to simultaneously degrade CP and carbofuran. To achieve this purpose, CP/carbofuran hydrolase genes and gfp were integrated into the chromosome of a biosafety strain Pseudomonas putida KT2440 using a chromosomal scarless modification strategy with upp as a counter-selectable marker...
July 15, 2016: Microbial Biotechnology
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