REHUNT: a reliable and open source package for restriction enzyme hunting.

BACKGROUND: Restriction enzymes are used frequently in biotechnology. However, manual mining of restriction enzymes is challenging. Furthermore, integrating available restriction enzymes into different bioinformatics systems is necessary for many biotechnological applications, such as polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Thus, in the present study, we developed the package REHUNT (Restriction Enzymes HUNTing), which mines restriction enzymes from the public database REBASE using a series of search operations. RESULTS: REHUNT is a reliable and open source package implemented in JAVA. It provides useful methods and manipulations for biological sequence analysis centered around restriction enzymes contained in REBASE. All available restriction enzymes for the imported biological sequences can be identified by REHUNT. Different genotypes can be identified using PCR-RFLP based on REHUNT for single nucleotide polymorphism (SNP), mutations, and the other variations. REHUNT robustly recognizes multiple inputs with different formats, e.g. regular DNA sequences, variation-in-sequence indicated by IUPAC code, as well as variation-in-sequence indicated by dNTPs format. Variations including di-, tri-, and tetra-allelic types and indel formats are also acceptable. Furthermore, REHUNT provides classified restriction enzymes output, including IUPAC and general sequence types, as well as commercial and non-commercial availabilities. REHUNT also enables analysis for high throughput screening (HTS) technologies. CONCLUSIONS: REHUNT is open source software with GPL v3 license and can be run on all platforms. Its features include: 1) Quick restriction enzymes search throughout a sequence based on the Boyer-Moore algorithm; 2) all available restriction enzymes provided and regularly updated from REBASE; 3) an open source API available of integrating all types of bioinformatics systems and applications; 4) SNP genotyping available for plant and animal marker-assisted breeding, and for human genetics; and 5) high throughput analysis available for Next Generation Sequencing (NGS). REHUNT not only to effectively looks for restriction enzymes in a sequence, but also available for SNP genotyping. Furthermore, it can be integrated into other biological and medical applications. REHUNT offers a convenient and flexible package for powerful restriction enzymes analyses in association studies, and supports high throughput analysis. The source codes and complete API documents are available at SourceForge: https://sourceforge.net/projects/rehunt/ , GitHub: https://github.com/yuhuei/rehunt , and at: https://sites.google.com/site/yhcheng1981/rehunt .

Design of Customize Interbody Fusion Cages of Ti64ELI with Gradient Porosity by Selective Laser Melting Process.

Intervertebral fusion surgery for spinal trauma, degeneration, and deformity correction is a major vertebral reconstruction operation. For most cages, the stiffness of the cage is high enough to cause stress concentration, leading to a stress shielding effect between the vertebral bones and the cages. The stress shielding effect affects the outcome after the reconstruction surgery, easily causing damage and leading to a higher risk of reoperation. A porous structure for the spinal fusion cage can effectively reduce the stiffness to obtain more comparative strength for the surrounding tissue. In this study, an intervertebral cage with a porous gradation structure was designed for Ti64ELI alloy powders bonded by the selective laser melting (SLM) process. The medical imaging software InVesalius and 3D surface reconstruction software Geomagic Studio 12 (Raindrop Geomagic Inc., Morrisville, NC, USA) were utilized to establish the vertebra model, and ANSYS Workbench 16 (Ansys Inc., Canonsburg, PA, USA) simulation software was used to simulate the stress and strain of the motions including vertical body-weighted compression, flexion, extension, lateral bending, and rotation. The intervertebral cage with a hollow cylinder had porosity values of 80-70-60-70-80% (from center to both top side and bottom side) and had porosity values of 60-70-80 (from outside to inside). In addition, according to the contact areas between the vertebras and cages, the shape of the cages can be custom-designed. The cages underwent fatigue tests by following ASTM F2077-17. Then, mechanical property simulations of the cages were conducted for a comparison with the commercially available cages from three companies: Zimmer (Zimmer Biomet Holdings, Inc., Warsaw, IN, USA), Ulrich (Germany), and B. Braun (Germany). The results show that the stress and strain distribution of the cages are consistent with the ones of human bone, and show a uniform stress distribution, which can reduce stress concentration.

Effects of Electropolishing on Mechanical Properties and Bio-Corrosion of Ti6Al4V Fabricated by Electron Beam Melting Additive Manufacturing.

Electron beam melting (EBM) has become one of the most promising additive manufacturing (AM) technologies. However, EBM tends to result in products with rougher surfaces due to the melt pool which causes adjacent powder particles to be sintered to the surface without being melted. Hence, it is necessary to improve the surface quality by post processing. The current study evaluates the tensile response of Ti6Al4V EBMed samples subject to various electropolishing (EP) treatments. The surface roughness Ra readings can be improved from over 24 µm down to about 4.5 µm by proper EP, resulting in apparent tensile elongation improvement from 7.6% to 11.6%, or a tensile plasticity increment of 53%, without any loss of elastic modulus or tensile strength. Moreover, the in-vitro bio-corrosion test in simulating body fluid (SBF) of the as-EBMed and EP-processed samples is also conducted. The potentiodynamic polarization reveals that the bio-corrosion resistance is improved by the lower Ra through proper EP treatments. This is due to the formation of a denser and more completely passivated oxide layer with less defects after proper EP duration. But when the EBMed samples are over-electropolished, nano pitting would induce a degraded bio-corrosion performance.

Improved Bioactivity of 3D Printed Porous Titanium Alloy Scaffold with Chitosan/Magnesium-Calcium Silicate Composite for Orthopaedic Applications.

Recently, cases of bone defects have been increasing incrementally. Thus, repair or replacement of bone defects is gradually becoming a huge problem for orthopaedic surgeons. Three-dimensional (3D) scaffolds have since emerged as a potential candidate for bone replacement, of which titanium (Ti) alloys are one of the most promising candidates among the metal alloys due to their low cytotoxicity and mechanical properties. However, bioactivity remains a problem for metal alloys, which can be enhanced using simple immersion techniques to coat bioactive compounds onto the surface of Ti⁻6Al⁻4V scaffolds. In our study, we fabricated magnesium-calcium silicate (Mg⁻CS) and chitosan (CH) compounds onto Ti⁻6Al⁻4V scaffolds. Characterization of these surface-modified scaffolds involved an assessment of physicochemical properties as well as mechanical testing. Adhesion, proliferation, and growth of human Wharton's Jelly mesenchymal stem cells (WJMSCs) were assessed in vitro. In addition, the cell attachment morphology was examined using scanning electron microscopy to assess adhesion qualities. Osteogenic and mineralization assays were conducted to assess osteogenic expression. In conclusion, the Mg⁻CS/CH coated Ti⁻6Al⁻4V scaffolds were able to exhibit and retain pore sizes and their original morphologies and architectures, which significantly affected subsequent hard tissue regeneration. In addition, the surface was shown to be hydrophilic after modification and showed mechanical strength comparable to natural bone. Not only were our modified scaffolds able to match the mechanical properties of natural bone, it was also found that such modifications enhanced cellular behavior such as adhesion, proliferation, and differentiation, which led to enhanced osteogenesis and mineralization downstream. In vivo results indicated that Mg⁻CS/CH coated Ti⁻6Al⁻4V enhances the bone regeneration and ingrowth at the critical size bone defects of rabbits. These results indicated that the proposed Mg⁻CS/CH coated Ti⁻6Al⁻4V scaffolds exhibited a favorable, inducive micro-environment that could serve as a promising modification for future bone tissue engineering scaffolds.

Bioprinting of artificial blood vessels.

Vascular networks have an important role to play in transporting nutrients, oxygen, metabolic wastes and maintenance of homeostasis. Bioprinting is a promising technology as it is able to fabricate complex, specific multi-cellular constructs with precision. In addition, current technology allows precise depositions of individual cells, growth factors and biochemical signals to enhance vascular growth. Fabrication of vascularized constructs has remained as a main challenge till date but it is deemed as an important stepping stone to bring organ engineering to a higher level. However, with the ever advancing bioprinting technology and knowledge of biomaterials, it is expected that bioprinting can be a viable solution for this problem. This article presents an overview of the biofabrication of vascular and vascularized constructs, the different techniques used in vascular engineering such as extrusion-based, droplet-based and laser-based bioprinting techniques, and the future prospects of bioprinting of artificial blood vessels.

Effect of Heat Treatment on Repetitively Scanned SLM NiTi Shape Memory Alloy.

Selective Laser Melting (SLM) has been implemented to address the difficulties in manufacturing complex nickel titanium (NiTi) structures. However, the SLM production of NiTi is much more challenging than the fabrication of conventional metals. Other than the need to have a high density that leads to excellent mechanical properties, strict chemical compositional control is required as well for the SLM NiTi parts to exhibit desirable phase transformation characteristics. In addition, acquiring a high transformation strain from the produced specimens is another challenging task. In the prior research, a new approach-repetitive scanning-was implemented to achieve these objectives. The repetitively scanned samples demonstrated an average of 4.61% transformation strain when subjected to the tensile test. Nevertheless, there is still room for improvement as the conventionally-produced NiTi can exhibit a transformation strain of about 6%. Hence, post-process heat treatment was introduced to improve the shape memory properties of the samples. The results showed an improvement when the samples were heat treated at a temperature of 400 °C for a period of 5 min. The enhancement in the shape memory behavior of the repetitively scanned samples was mainly attributed to the formation of fine Ni4Ti3 metastable precipitates.

A Review of Selective Laser Melted NiTi Shape Memory Alloy.

NiTi shape memory alloys (SMAs) have the best combination of properties among the different SMAs. However, the limitations of conventional manufacturing processes and the poor manufacturability of NiTi have critically limited its full potential applicability. Thus, additive manufacturing, commonly known as 3D printing, has the potential to be a solution in fabricating complex NiTi smart structures. Recently, a number of studies on Selective Laser Melting (SLM) of NiTi were conducted to explore the various aspects of SLM-produced NiTi. Compared to producing conventional metals through the SLM process, the fabrication of NiTi SMA is much more challenging. Not only do the produced parts require a high density that leads to good mechanical properties, strict composition control is needed as well for the SLM NiTi to possess suitable phase transformation characteristics. Additionally, obtaining a good shape memory effect from the SLM NiTi samples is another challenging task that requires further understanding. This paper presents the results of the effects of energy density and SLM process parameters on the properties of SLM NiTi. Its shape memory properties and potential applications were then reviewed and discussed.

Effective Natural PCR-RFLP Primer Design for SNP Genotyping Using Teaching-Learning-Based Optimization With Elite Strategy.

SNP (single nucleotide polymorphism) genotyping is the determination of genetic variations of SNPs between members of a species. In many laboratories, PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) is a usually used biotechnology for SNP genotyping, especially in small-scale basic research studies of complex genetic diseases. PCR-RFLP requires an available restriction enzyme at least for identify a target SNP and an effective primer pair conforms numerous constraints. However, the lots of restriction enzymes, tedious sequence and complicated constraints make the mining of available restriction enzymes and the design of effective primer pairs become a major challenge. In the study, we propose a novel and available CI (Computation Intelligence)-based method called TLBO (teaching-learning-based optimization) and introduce the elite strategy to design effective primer pairs. Three common melting temperature computations are available in the method. REHUNT (Restriction Enzymes HUNTing) is first combined with the method to mine available restriction enzymes. Robust in silico simulations for the GA (genetic algorithm), the PSO (particle swarm optimization), and the method for natural PCR-RFLP primer design in the SLC6A4 gene with two hundred and eighty-eight SNPs had been performed and compared. These methods had been implemented in JAVA and they are freely available at https://sites.google.com/site/yhcheng1981/tlbonpd-elite for users of academic and non-commercial interests.