Killing Strategies Research Articles

Antimicrobial polymeric coatings synthesized by solvent-free initiated Chemical Vapor Deposition: A review

Microbial adhesion on surfaces may cause infectious diseases that seriously threaten human health, which can be controlled by antimicrobial surfaces. Antimicrobial polymeric coatings can endow surfaces with biocidal properties to address infectious diseases. Initiated Chemical Vapor Deposition (iCVD) is a novel technique to synthesize high-quality polymeric antimicrobial coatings on various surfaces. Owing to the solvent-free and mild deposition conditions, iCVD facilitates solvent-susceptible, delicate, or fragile substrates. Herein, we provided a comprehensive review of antimicrobial coatings fabricated by iCVD. First, iCVD was introduced. Then, antimicrobial iCVD coatings were discussed based on killing and antifouling strategies. The microbe-killing coatings were discussed according to release-killing and contact-killing approaches. Next, hydrophilic, hydrophobic, and amphiphilic antifouling coatings were reviewed. Finally, micro/nano-structured antimicrobial surfaces fabricated via iCVD were also discussed, followed by the future directions of iCVD coatings in the antimicrobial field.

CRISPR-Cas9 for selective targeting of somatic mutations in pancreatic cancers.

Somatic mutations are desirable targets for selective elimination of cancer, yet most are found within noncoding regions. We have adapted the CRISPR-Cas9 gene editing tool as a novel, cancer-specific killing strategy by targeting the subset of somatic mutations that create protospacer adjacent motifs (PAMs), which have evolutionally allowed bacterial cells to distinguish between self and non-self DNA for Cas9-induced double strand breaks. Whole genome sequencing (WGS) of paired tumor minus normal (T-N) samples from three pancreatic cancer patients (Panc480, Panc504, and Panc1002) showed an average of 417 somatic PAMs per tumor produced from single base substitutions. Further analyses of 591 paired T-N samples from The International Cancer Genome Consortium found medians of ∼455 somatic PAMs per tumor in pancreatic, ∼2800 in lung, and ∼3200 in esophageal cancer cohorts. Finally, we demonstrated 69-99% selective cell death of three targeted pancreatic cancer cell lines using 4-9 sgRNAs designed using the somatic PAM discovery approach. We also showed no off-target activity from these tumor-specific sgRNAs in either the patient's normal cells or an irrelevant cancer using WGS. This study demonstrates the potential of CRISPR-Cas9 as a novel and selective anti-cancer strategy, and supports the genetic targeting of adult cancers.

First person – Fabrizia Zevolini

ABSTRACT First Person is a series of interviews with the first authors of a selection of papers published in Journal of Cell Science, helping researchers promote themselves alongside their papers. Fabrizia Zevolini is first author on ‘ Polo-like kinase 1 regulates immune synapse assembly and cytotoxic T cell function by driving microtubule dynamics’, published in JCS. Fabrizia is a postdoc in the lab of Cosima T. Baldari at the University of Siena, Italy, investigating cytotoxic T lymphocyte effector functions, with a focus on their killing apparatus and strategies.

Eradication of human immunodeficiency virus-1 reservoir in the brain microglia.

Eradication of human immunodeficiency virus-1 reservoir in the brain microglia.

Dynamic alterations in PD-1/PD-L1 expression level and immune cell profiles based on radiation response status in mouse tumor model.

Based on the immunologic effects of anti-cancer treatment and their therapeutic implications, we evaluated radiotherapy (RT)-induced dynamic alterations in programmed death-1 (PD-1)/PD ligand-1 (PD-L1) expression profiles. Local RT with 2 Gy × 5 or 7.5 Gy × 1 was administered to the CT26 mouse model. Thereafter, tumors were resected and evaluated at the following predefined timepoints according to radiation response status: baseline, early (immediately after RT), middle (beginning of tumor shrinkage), late (stable status with RT effect), and progression (tumor regrowth). PD-1/PD-L1 activity and related immune cell profiles were quantitatively assessed. RT upregulated PD-L1 expression in tumor cells from the middle to late phase; however, the levels subsequently decreased to levels comparable to baseline in the progression phase. RT with 2 Gy × 5 induced a higher frequency of PD-L1+ myeloid-derived suppressor cells, with a lesser degree of tumor regression, compared to 7.5 Gy. The proportion of PD-1+ and interferon (IFN)-γ+CD8α T cells continued to increase. The frequency of splenic PD-1+CD8+ T cells was markedly elevated, and was sustained longer with 2 Gy × 5. Based on the transcriptomic data, RT stimulated the transcription of immune-related genes, leading to sequentially altered patterns. The dynamic alterations in PD-1/PD-L1 expression level were observed according to the time phases of tumor regression. This study suggests the influence of tumor cell killing and radiation dosing strategy on the tumor immune microenvironment.

TCR-mimic bispecific antibodies to target the HIV-1 reservoir

HIV-1 infection is incurable due to the persistence of the virus in a latent reservoir of resting memory CD4+ T cells. “Shock-and-kill” approaches that seek to induce HIV-1 gene expression, protein production, and subsequent targeting by the host immune system have been unsuccessful due to a lack of effective latency-reversing agents (LRAs) and kill strategies. In an effort to develop reagents that could be used to promote killing of infected cells, we constructed T cell receptor (TCR)-mimic antibodies to HIV-1 peptide-major histocompatibility complexes (pMHC). Using phage display, we panned for phages expressing antibody-like variable sequences that bound HIV-1 pMHC generated using the common HLA-A*02:01 allele. We targeted three epitopes in Gag and reverse transcriptase identified and quantified via Poisson detection mass spectrometry from cells infected in vitro with a pseudotyped HIV-1 reporter virus (NL4.3 dEnv). Sequences isolated from phages that bound these pMHC were cloned into a single-chain diabody backbone (scDb) sequence, such that one fragment is specific for an HIV-1 pMHC and the other fragment binds to CD3ε, an essential signal transduction subunit of the TCR. Thus, these antibodies utilize the sensitivity of T cell signaling as readouts for antigen processing and as agents to promote killing of infected cells. Notably, these scDbs are exquisitely sensitive and specific for the peptide portion of the pMHC. Most importantly, one scDb caused killing of infected cells presenting a naturally processed target pMHC. This work lays the foundation for a novel therapeutic killing strategy toward elimination of the HIV-1 reservoir.

Cell-to-cell variability in inducible Caspase9-mediated cell death

iCasp9 suicide gene has been widely used as a promising killing strategy in various cell therapies. However, different cells show significant heterogeneity in response to apoptosis inducer, posing challenges in clinical applications of killing strategy. The cause of the heterogeneity remains elusive so far. Here, by simultaneously monitoring the dynamics of iCasp9 dimerization, Caspase3 activation, and cell fate in single cells, we found that the heterogeneity was mainly due to cell-to-cell variability in initial iCasp9 expression and XIAP/Caspase3 ratio. Moreover, multiple-round drugging cannot increase the killing efficiency. Instead, it will place selective pressure on protein levels, especially on the level of initial iCasp9, leading to drug resistance. We further show this resistance can be largely eliminated by combinatorial drugging with XIAP inhibitor at the end, but not at the beginning, of the multiple-round treatments. Our results unveil the source of cell fate heterogeneity and drug resistance in iCasp9-mediated cell death, which may enlighten better therapeutic strategies for optimized killing.

Tumor targeting and penetrating biomimetic mesoporous polydopamine nanoparticles facilitate photothermal killing and autophagy blocking for synergistic tumor ablation

The synergistic manipulation of autophagy blocking with tumor targeting and penetration effects to enhance cancer cell killing during photothermal therapy (PTT) remains a substantial challenge. Herein, we fabricated a biomimetic nanoplatform by precisely coating homologous prostate cancer cell membranes (CMs) onto the surface of mesoporous polydopamine nanoparticles (mPDA NPs) encapsulating the autophagy inhibitor chloroquine (CQ) for synergistically manipulating PTT and autophagy for anticancer treatment. The resulting biomimetic mPDA@CMs NPs-CQ system could escape macrophage phagocytosis, overcome the vascular barrier, and home in the homologous prostate tumor xenograft with high tumor targeting and penetrating efficiency. The mPDA NPs core endowed the mPDA@CMs NPs-CQ with good photothermal capability to mediate PTT killing of prostate cancer cells, while NIR-triggered CQ release from the nanosystem further arrested PTT-induced protective autophagy of cancer cells, thus weakening the resistance of prostate cancer cells to PTT. This combined PTT killing and autophagy blocking anticancer strategy could induce significant autophagosome accumulation, ROS generation, mitochondrial damage, endoplasmic reticulum stress, and apoptotic signal transduction, which finally results in synergistic prostate tumor ablation in vivo. This prostate cancer biomimetic nanosystem with synergistically enhanced anticancer efficiency achieved by manipulating PTT killing and autophagy blocking is expected to serve as a more effective anticancer strategy against prostate cancer. Statement of significanceAutophagy is considered as one of the most efficient rescuer and reinforcement mechanisms of cancer cells against photothermal therapy (PTT)-induced cancer cell eradication. How to synergistically manipulate autophagy blocking with significant tumor targeting and penetration to enhance PTT-mediated cancer cell killing remains a substantial challenge. Herein, we fabricated a biomimetic nanoplatform by precisely coating homologous cancer cell membranes onto the surface of mesoporous polydopamine nanoparticles with encapsulation of the autophagy inhibitor chloroquine for synergistic antitumor treatment with high tumor targeting and penetrating efficiency both in vitro and in vivo. This biomimetic nanosystem with synergistically enhanced anticancer efficiency by manipulating PTT killing and autophagy blocking is expected to serve as a more effective anticancer strategy.

Moniliophthora perniciosa, the Causal Agent of Cacao Witches’ Broom Disease Is Killed in vitro by Saccharomyces cerevisiae and Wickerhamomyces anomalus Yeasts

Cacao plantations from South America have been afflicted with the severe fungal disease known as Witches’ Broom Disease (WBD), caused by the basidiomycete Moniliophthora perniciosa. Yeasts are increasingly recognized as good fungal biocides, although their application is still mostly restricted to the postharvest control of plant and fruit decay. Their possible utilization in the field, in a preharvest phase, is nevertheless promising, particularly if the strains are locally adapted and evolved and if they belong to species considered safe for man and the environment. In this work, a group of yeast strains originating from sugarcane-based fermentative processes in Brazil, the cacao-producing country where the disease is most severe, were tested for their ability to antagonize M. perniciosa in vitro. Wickerhamomyces anomalus LBCM1105 and Saccharomyces cerevisiae strains LBCM1112 from spontaneous fermentations used to produce cachaça, and PE2 widely used in Brazil in the industrial production of bioethanol, efficiently antagonized six strains of M. perniciosa, originating from several South American countries. The two fastest growing fungal strains, both originating from Brazil, were further used to assess the mechanisms underlying the yeasts’ antagonism. Yeasts were able to inhibit fungal growth and kill the fungus at three different temperatures, under starvation, at different culture stages, or using an inoculum from old yeast cultures. Moreover, SEM analysis revealed that W. anomalus and S. cerevisiae PE2 cluster and adhere to the hyphae, push their surface, and fuse to them, ultimately draining the cells. This behavior concurs with that classified as necrotrophic parasitism/mycoparasitism. In particular, W. anomalus within the adhered clusters appear to be ligated to each other through roundish groups of fimbriae-like structures filled with bundles of microtubule-sized formations, which appear to close after cells detach, leaving a scar. SEM also revealed the formation of tube-like structures apparently connecting yeast to hypha. This evidence suggests W. anomalus cells form a network of yeast cells connecting with each other and with hyphae, supporting a possible cooperative collective killing and feeding strategy. The present results provide an initial step toward the formulation of a new eco-friendly and effective alternative for controlling cacao WBD using live yeast biocides.

The recent progress on metal-organic frameworks for phototherapy.

Some infectious or malignant diseases such as cancers are seriously threatening the health of human beings all over the world. The commonly used antibiotic therapy cannot effectively treat these diseases within a short time, and also bring about adverse effects such as drug resistance and immune system damage during long-term systemic treatment. Phototherapy is an emerging antibiotic-free strategy to treat these diseases. Upon light irradiation, phototherapeutic agents can generate cytotoxic reactive oxygen species (ROS) or induce a temperature increase, which leads to the death of targeted cells. These two kinds of killing strategies are referred to as photodynamic therapy (PDT) and photothermal therapy (PTT), respectively. So far, many photo-responsive agents have been developed. Among them, the metal-organic framework (MOF) is becoming one of the most promising photo-responsive materials because its structure and chemical compositions can be easily modulated to achieve specific functions. MOFs can have intrinsic photodynamic or photothermal ability under the rational design of MOF construction, or serve as the carrier of therapeutic agents, owing to its tunable porosity. MOFs also provide feasibility for various combined therapies and targeting methods, which improves the efficiency of phototherapy. In this review, we firstly investigated the principles of phototherapy, and comprehensively summarized recent advances of MOF in PDT, PTT and synergistic therapy, from construction to modification. We expect that our demonstration will shed light on the future development of this field, and bring it one step closer to clinical trials.

Morphological convergence obscures functional diversity in sabre-toothed carnivores.

The acquisition of elongated, sabre-like canines in multiple vertebrate clades during the last 265 Myr represents a remarkable example for convergent evolution. Due to striking superficial similarities in the cranial skeleton, the same or similar skull and jaw functions have been inferred for sabre-toothed species and interpreted as an adaptation to subdue large-bodied prey. However, although some sabre-tooth lineages have been classified into different ecomorphs (dirk-tooths and scimitar-tooths) the functional diversity within and between groups and the evolutionary paths leading to these specializations are unknown. Here, we use a suite of biomechanical simulations to analyse key functional parameters (mandibular gape angle, bending strength, bite force) to compare the functional performance of different groups and to quantify evolutionary rates across sabre-tooth vertebrates. Our results demonstrate a remarkably high functional diversity between sabre-tooth lineages and that different cranial function and prey killing strategies evolved within clades. Moreover, different biomechanical adaptations in coexisting sabre-tooth species further suggest that this functional diversity was at least partially driven by niche partitioning.

Effect of different killing methods on physicochemical traits, nutritional characteristics, in vitro human digestibility and oxidative stability during storage of the house cricket (Acheta domesticus L.)

The present research studied the effect of different killing methods on various quality attributes of the house cricket (Acheta domesticus) powder: physicochemical traits (pH, colour, radical scavenging activity, browning index, protein oxidation), nutritional profile (proximate composition), in vitro human digestibility, and oxidative stability of lipids during storage. With this purpose, seven different killing methods were compared: blanching (40 s at 100 °C), steaming (2 min 15 s), freezing (2 h 10 min at −20 °C), carbon dioxide (2 h 7 min at 25 °C), vacuum (1 h 35 min at 25 °C), plastic bag (3 h 1 min at 25 °C), and carbon dioxide + blanching (40 min carbon dioxide at 25 °C + 40 s blanching). All killing methods reduced the radical scavenging activity (p < 0.001) and increased the browning index (p < 0.001) of the cricket powder. Overall, cricket powder obtained through carbon dioxide, vacuum and plastic bag killing methods provided the best proximate composition (p < 0.001). Freezing, carbon dioxide, vacuum and plastic bag provided cricket powder with the highest polyunsaturated fatty acids proportion, thus having the healthiest lipid profile (p < 0.001), and guaranteed the lowest oxidative degree of lipids (p < 0.001). In the same groups, however, the lowest in vitro human digestibility was also observed (p < 0.001). Overall, none of the tested methods had detrimental effects on product quality and, at present, the choice of the most appropriate killing strategy seems a matter of equipment availability and cost-efficiency evaluations. Industrial relevanceThe present study provides new insights on the impact of the killing method on the final quality of cricket powder, a versatile, nutrient-rich food. The choice of the most appropriate method, together with its industrial implementation, are fundamental aspects to guarantee satisfactory quality standards and consumer's health.

Analysis of G-Queue with Pseudo-Fault and Multiple Working Vacations

This paper presents a new model of discrete time Geo/Geo/1 repairable queueing system with pseudo-fault, negative customers and multiple working vacations. The authors assume that system service may be interrupted by breakdown or pseudo-fault, this system may become disabled only when it is in a regular busy period, and negative customers adopt two types of typical killing strategies. In this paper, the authors know that the evolution of the system can be described by a two-dimensional Markov chain, and the two-dimensional Markov chain satisfies the condition of quasi birth and death chains. Based on the method of matrix-geometric solution, the authors obtain distributions for the stationary queue length in RCH and RCE strategy, respectively. Moreover, the reliability of the system is analyzed and the number of customers and waiting time of a customer in the system in steady state are obtained. The authors analyze the impact of two killing strategies on the system comparatively. This paper studies the individually and socially optimal behaviors of positive customers, and presents a pricing policy for positive customers, therefore, the authors obtain the socially optimal arrival rate. Various numerical results are provided to show the change of performance measures.

Black soldier fly (Hermetia illucens L.): effect on the fat integrity using different approaches to the killing of the prepupae

The increase of waste, due to the rise of the world population, renders the use of the black soldier fly (BSF;Hermetia illucens) as agri-food leftover biotransformer very attractive. Indeed, feeding on these substrates, BSF is capable of transforming them into valuable fat, proteins, and chitin. The present study is aimed at evaluating different approaches to the killing of the prepupae in order to assess which is associated with the lowest production of artefacts, notably free fatty acids. Folch extraction method, via an Ultra-Turrax homogeniser, was selected to isolate and analyse prepupal fat. The same method was also applied for a direct grinding method to kill black soldier fly prepupae. The integrity of fat originated from this direct grinding approach to the killing was compared in terms of fatty acid profile with the samples obtained with two other killing procedures, i.e. freezing and blanching. Direct grinding proved to be a better preservative of fat integrity in terms of lowest amount of free fatty acids (about 2.5%) in comparison with freezing (about 15%), but also with blanching (about 10%). Furthermore, the direct grinding is a killing strategy that reduces energetic demand and process time and helps lower analytical costs.

Skeleton of a new owl from the early Eocene of North America (Aves, Strigiformes) with an accipitrid-like foot morphology

ABSTRACT We describe a partial skeleton of a large-sized owl from Wasatchian strata of the Willwood Formation (Wyoming, U.S.A.). The holotype of Primoptynx poliotauros, gen. et sp. nov., includes all major postcranial bones and is one of the most substantial Paleogene records of the Strigiformes. The fossil shows that owls exhibited a considerable morphological diversity in the early Eocene of North America and occupied disparate ecological niches. As in the protostrigid taxon Minerva from the late early to early middle Eocene of North America, but unlike in extant owls, the ungual phalanges of the hallux and the second toe of the new species are distinctly larger than those of the other toes. Primoptynx poliotauros gen. et sp. nov., however, does not exhibit the derived tibiotarsus morphology of the Protostrigidae. Even though the new species may well be a stem group representative of protostrigid owls, current data do not allow an unambiguous phylogenetic placement. Concerning the size of the ungual phalanges, the feet of P. poliotauros correspond to those of extant hawks and allies (Accipitridae). We therefore hypothesize that it used its feet to dispatch prey items in a hawk-like manner, whereas extant owls kill prey with their beak. Primoptynx and protostrigid owls were possibly specialized in foraging on prey items that required an accipitrid-like killing strategy, such as larger-sized or more defensive mammals. The extinction of these peculiar owls may have been related to the radiation of accipitrid diurnal birds of prey, which appear to have diversified in the late Eocene and early Oligocene.

Identification and Characterization of Nematicidal Volatile Organic Compounds from Deep-Sea Virgibacillus dokdonensis MCCC 1A00493.

Root-knot nematode diseases cause severe yield and economic losses each year in global agricultural production. Virgibacillus dokdonensis MCCC 1A00493, a deep-sea bacterium, shows a significant nematicidal activity against Meloidogyne incognita in vitro. However, information about the active substances of V. dokdonensis MCCC 1A00493 is limited. In this study, volatile organic compounds (VOCs) from V. dokdonensis MCCC 1A00493 were isolated and analyzed through solid-phase microextraction and gas chromatography–mass spectrometry. Four VOCs, namely, acetaldehyde, dimethyl disulfide, ethylbenzene, and 2-butanone, were identified, and their nematicidal activities were evaluated. The four VOCs had a variety of active modes on M. incognita juveniles. Acetaldehyde had direct contact killing, fumigation, and attraction activities; dimethyl disulfide had direct contact killing and attraction activities; ethylbenzene had an attraction activity; and 2-butanone had a repellent activity. Only acetaldehyde had a fumigant activity to inhibit egg hatching. Combining this fumigant activity against eggs and juveniles could be an effective strategy to control the different developmental stages of M. incognita. The combination of direct contact and attraction activities could also establish trapping and killing strategies against root-knot nematodes. Considering all nematicidal modes or strategies, we could use V. dokdonensis MCCC 1A00493 to set up an integrated strategy to control root-knot nematodes.

DNA computing inspired deep networks design

Deep neural networks have gained state-of-the-art results in many applications, such as pattern recognition and computer vision. However, most of the deep neural networks are designed manually by researchers. This architecture design process is generally time consuming and needs much expertise. Hence, automatic neural network design becomes an important issue. In this paper, we propose a novel method, called DNA computing inspired networks design (DNAND), to automatically learn high performance deep networks. In DNAND, we use DNA strands to represent blocks of a model, and these DNA strands are reacted to construct the overall networks according to the base pairing principle. We also present the killing strategy, with which we stop training “bad” models if they fail to reach the specific accuracy threshold on the validation set, so as to reduce the computational cost and accelerate the learning process. Extensive experiments on image classification and detection data sets demonstrate the effectiveness of the proposed method.

Non-mimetic shiny cowbird nestlings escape discrimination by baywings in absence of host nest mates

Nestlings of obligate brood parasites must obtain resources from heterospecific hosts that are attuned to the solicitation behaviours of their own progeny. Failing to match the appropriate stimuli may result in suboptimal provisioning or even the starvation of parasite young. Parasitic nestlings could overcome it by sharing the nest with host young, as long as they are able to compete efficiently for parental feedings. Here, we examined if non-mimetic shiny cowbird (Molothrus bonariensis) nestlings reared alone fail to elicit sufficient parental care from the grayish baywing (Agelaioides badius), a host that discriminates between their own and parasitic nestlings based on species-specific begging cues. We manipulated baywing broods to assess the ability of shiny cowbird nestlings to elicit parental provisioning and survive without host nest mates. Host provisioning rates and survival did not differ between shiny cowbirds reared alone and host nestlings in singleton broods. Also, growth patterns of experimental nestlings reared alone were similar to those reported for shiny cowbirds reared alongside baywing young. Hence, we did not find support for the hypothesis that non-mimetic nestlings require the assistance of host nest mates to elicit sufficient parental care from its baywing host. Furthermore, we found that shiny cowbirds that were reared alone continued receiving care from baywings after fledging, unlike shiny cowbirds from mixed broods. Our results add evidence to the idea of opposing selective pressures on the evolution of nest mate acceptance in cowbirds and suggest context-dependent host’s discrimination abilities that deserve further investigation. The evolution of nest mate killing versus tolerance strategies is a long-standing question in the study of brood parasitism. Parasitic nestlings may benefit from sharing the nest with host young if they collectively induce more parental provisioning and the parasite can benefit from it. Host nest mates could also confer protection against discrimination when hosts respond preferentially to stimuli of their own kind. We examined this idea in a generalist brood parasite, the shiny cowbird (Molothrus bonariensis), and a host able to discriminate species-specific begging cues. The results showed that shiny cowbirds reared alone performed well and circumvent host discrimination also after leaving the nest. Our study supports the existence of trade-offs on the evolution of nest mate killing behaviours and adds evidence that context-dependent host’s discrimination abilities similar to those observed towards parasitic eggs can be at play towards parasitic fledglings.

THE US STRATEGY OF ARMED DRONE USAGE IN FIGHTING THE TALIBAN ELEMENTS IN PAKISTAN (2004-2016)

&lt;p&gt;Prior to the Trump’s presidency the US have performed as many as 400 armed drone strikes in Pakistan, killing approximately 3.000 terrorist targets including key leaders that hold important value to the organizations. More than a decade these attacks were performed, yet we have not seen the end of the terrorist and insurgent’s presence in Pakistan. This leads into a thought whether the attacks were effective or not. Therefore, this study aims to analyze, predominantly evaluating the use of the targeted killing strategy in this US counterterrorism effort. The study uses qualitative research method by using Robert A. Pape’s perspective in strategic effectiveness and David Galula’s counterinsurgency perspective. Those perspectives pointed out the cause of this situation is due to the inability of the attacked target set to trigger the destruction of the movement, the inability of the US to secure the ends, ways, and means of the operation which involve the dissociation between the population and the insurgents, their ability to forge proper cooperation with its local ally, and the un-favoring view of the local population toward its targeted killings practice has resulted in the infectivity of the strategy and a prolonged fight between the US and its opponent. In sum, it can be inferred that despite capable of eliminating numerous high-value targets the targeted killing strategy is deemed ineffective due to its inability to procure full destruction of its opponent and due to the inability of the US to secure the ends, ways, and means of the operation which would have increase the success of the strategy.&lt;/p&gt;

Human T-cell receptor V gene segment of alpha and beta families: A revised primer design strategy.

The application of human TCR in cancer immunotherapy has gained momentum with developments in tumor killing strategies using endogenous adaptive immune responses. The successful coverage of a diverse TCR repertoire is mainly attributed to the primer design of the human TCR V genes. Here, we present a refined primer design strategy of the human TCR V gene by clustering V gene sequence homolog for degenerate primer design based on the data from IMGT. The primers designed were analyzed and the PCR efficiency of each primer set was optimized. A total of 112 alpha and 160 beta sequences were aligned and clustered using a phylogram yielding 32 and 27 V gene primers for the alpha and beta family. The new primer set was able to provide 93.75% and 95.63% coverage for the alpha and beta family, respectively. A semi-qualitative approach using the designed primer set was able to provide a relative view of the TCR V gene diversity in different populations. Taken together, the new primers provide a more comprehensive coverage of the TCR gene diversity for improved TCR library generation and TCR V gene analysis studies.