Sanger sequencing - a hands-on simulation

2018 ◽  
Author(s):  
Jared Young
Keyword(s):  
Sanger Sequencing ◽  
Hands On

scholarly journals A rapid, cost efficient and simple method to identify current SARS-CoV-2 variants of concern by Sanger sequencing part of the spike protein gene

2021 ◽  
Author(s):  
Tue Sparholt Joergensen ◽  
Kai Blin ◽  
Franziska Kuntke ◽  
Henrik K. Salling ◽  
Thomas Yssing Michaelsen ◽  
...  
Keyword(s):  
Sanger Sequencing ◽  
Turnaround Time ◽  
Contact Tracing ◽  
The Novel ◽  
Simple Method ◽  
Hands On ◽  
Sequencing Part ◽  
Cost Efficient ◽  
Set Up ◽  
Novel Coronavirus

In 2020, the novel coronavirus, SARS-CoV-2, caused a pandemic, which is still raging at the time of writing this. Many countries have set up high throughput RT-qPCR based diagnostics for people with COVID-19 symptoms and for the wider population. In addition, with the use of whole genome sequencing (WGS) new lineages of SARS-CoV-2 have been identified that have been associated with increased transmissibility or altered vaccine efficacy, so-called Variants of Concern (VoC). WGS is generally too labor intensive and expensive to be applied to all positive samples from the diagnostic tests, and often has a turnaround time too long to enable VoC focused contact tracing. Here, we propose to use Sanger sequencing for the detection of common variants of concern and key mutations in early 2021, using a single set of the recognized ARTIC Network primers. The proposed setup relies entirely on materials and methods already in use in diagnostic RT-qPCR labs and on existing infrastructure from companies that have specialized in cheap and rapid turnaround Sanger sequencing. In addition, we provide an automated mutation calling software (https://github.com/kblin/covid-spike-classification). We have validated the setup on 195 SARS-CoV-2 positive samples, and we were able to profile >85% of RT-qPCR positive samples, where the last 15% largely stem from samples with low viral count. At approximately 4 € per sample in material cost, with minimal hands-on time, little data handling, and a turnaround time of less than 30 hours, the setup is simple enough to be implemented in any SARS-CoV-2 RT-qPCR diagnostic lab. Our protocol provides results that can be used to focus contact-tracing efforts and it is cheap enough for the tracking and surveillance of all positive samples for emerging variants such as B.1.1.7, B.1.351 and P.1 as of January 2021.

scholarly journals AB12PHYLO: an integrated pipeline for Maximum Likelihood phylogenetic inference from ABI trace data

2021 ◽  
Author(s):  
Leo Kaindl ◽  
Corinn Small ◽  
Remco Stam
Keyword(s):  
Sanger Sequencing ◽  
Cost Effective ◽  
Reference Database ◽  
Sequencing Data ◽  
Type Specimens ◽  
Multiple Sequence ◽  
The Past ◽  
Hands On ◽  
High Degree ◽  
Command Line Version

AbstractMulti-gene phylogenies constructed from multiplexed and Sanger sequencing data are regularly used in mycology and other disciplines as a cost-effective way of species identification and as a first means to investigate genetic diversity samples.Today, a number of tools exist for each of the steps in this analysis, including quality control and trimming, the generation of a multiple sequence alignment (MSA), extraction of informative sites, and the construction of the final phylogenetic tree. A BLAST search in a reference database is often performed to identify sequences of type specimens to compare the samples with in the phylogeny. Made over the past decades, these tools are all independent from and often not perfectly adapted to one another.We present AB12PHYLO, an integrated pipeline that can perform all necessary steps from reading in raw Sanger sequencing data through visualizing and editing phylogenies. In addition, AB12PHYLO can calculate basic summary statistics for each gene in the phylogeny.AB12PHYLO is designed as a wrapper of several open access and commonly used tools for each of the intermediate stages, and intended to simplify the phylogenetic pipeline while still allowing a high degree of access. It comes as a command-line version for the highest reproducibility and an intuitive graphical user interface (GUI) for easy adoption by IT-agnostic end-users. The use of AB12PHYLO significantly reduces the hands-on working time for these analyses.

scholarly journals TnClone: high-throughput clonal analysis using Tn5-mediated library construction and de novo assembly

2018 ◽  
Author(s):  
Byungjin Hwang ◽  
Sunghoon Heo ◽  
Namjin Cho ◽  
Duhee Bang
Keyword(s):  
Sanger Sequencing ◽  
Large Scale ◽  
De Novo ◽  
Low Frequency ◽  
Clonal Analysis ◽  
Hands On ◽  
The Cost ◽  
Next Generation Sequencing Ngs ◽  
User Friendly ◽  
Analysis Platform

ABSTRACTA typical molecular cloning procedure requires Sanger sequencing for validation, which becomes cost-prohibitive and labour-intensive for large-scale clonal analysis of genotype-phenotype studies. Here we present a Tn5-mediated clonal analysis platform TnClone, which uses next-generation sequencing (NGS) to rapidly and cost-effectively analyze a large number of clones. We also developed a user-friendly graphical user interface and have provided general guidelines for conducting validation experiments. Using TnClone, we achieved more than 20-fold cost reduction compared with the cost incurred using conventional Sanger sequencing and detected low-frequency mutant clones (~10%) in mixed samples. We tested our programme and achieved 99.4% sensitivity. Our platform provides rapid turnaround with minimal hands-on time for secondary evaluation as NGS technology continues to evolve.

The Instructional SEM Laboratory at Iowa State University

1996 ◽  
Vol 54 ◽  
pp. 396-397
Author(s):  
L. S. Chumbley ◽  
M. Meyer ◽  
K. Fredrickson ◽  
F.C. Laabs
Keyword(s):  
Materials Science ◽  
Computer Network ◽  
Energy Dispersive Spectrometer ◽  
Computer Control ◽  
State University ◽  
Iowa State University ◽  
Internet Server ◽  
Schematic Layout ◽  
Hands On ◽  
Improve Instruction

The Materials Science Department at Iowa State University has developed a laboratory designed to improve instruction in the use of the scanning electron microscope (SEM). The laboratory makes use of a computer network and a series of remote workstations in a classroom setting to provide students with increased hands-on access to the SEM. The laboratory has also been equipped such that distance learning via the internet can be achieved.A view of the laboratory is shown in Figure 1. The laboratory consists of a JEOL 6100 SEM, a Macintosh Quadra computer that acts as a server for the network and controls the energy dispersive spectrometer (EDS), four Macintosh computers that act as remote workstations, and a fifth Macintosh that acts as an internet server. A schematic layout of the classroom is shown in Figure 2. The workstations are connected directly to the SEM to allow joystick and computer control of the microscope. An ethernet connection between the Quadra and the workstations allows students seated there to operate the EDS. Control of the microscope and joystick is passed between the workstations by a switch-box assembly that resides at the microscope console. When the switch-box assembly is activated a direct serial line is established between the specified workstation and the microscope via the SEM’s RS-232.

Students Involvement in Multilingual and Multicultural Community Services

2015 ◽  
Vol 22 (2) ◽  
pp. 77-89 ◽  
Author(s):  
Ying-Chiao Tsao
Keyword(s):  
Service Learning ◽  
Cultural Competence ◽  
Linguistically Diverse ◽  
Community Services ◽  
Cultural Humility ◽  
Self Assessment ◽  
Multicultural Community ◽  
Raising Awareness ◽  
Hands On ◽  
Teaching Learning

Promoting cultural competence in serving diverse clients has become critically important across disciplines. Yet, progress has been limited in raising awareness and sensitivity. Tervalon and Murray-Garcia (1998) believed that cultural competence can only be truly achieved through critical self-assessment, recognition of limits, and ongoing acquisition of knowledge (known as “cultural humility”). Teaching cultural humility, and the value associated with it remains a challenging task for many educators. Challenges inherent in such instruction stem from lack of resources/known strategies as well as learner and instructor readiness. Kirk (2007) further indicates that providing feedback on one's integrity could be threatening. In current study, both traditional classroom-based teaching pedagogy and hands-on community engagement were reviewed. To bridge a gap between academic teaching/learning and real world situations, the author proposed service learning as a means to teach cultural humility and empower students with confidence in serving clients from culturally/linguistically diverse backgrounds. To provide a class of 51 students with multicultural and multilingual community service experience, the author partnered with the Tzu-Chi Foundation (an international nonprofit organization). In this article, the results, strengths, and limitations of this service learning project are discussed.

A ’Hands-On’ Approach

ASHA Leader ◽  
2012 ◽  
Vol 17 (9) ◽  
pp. 55-55
Author(s):  
Kimberly Abts
Keyword(s):  
Hands On

Telepractice: A Survey of AuD Students Pre- and Post-Telepractice

2017 ◽  
Vol 2 (18) ◽  
pp. 28-41
Author(s):  
Kelli M. Watts ◽  
Laura B. Willis
Keyword(s):  
Doctoral Students ◽  
Graduate School ◽  
Professional Services ◽  
Hands On ◽  
Before And After ◽  
Lack Of Knowledge

Telepractice, defined by the American Speech-Language-Hearing Association (ASHA, n.d.) as “the application of telecommunications technology to the delivery of professional services at a distance by linking clinician to client, or clinician to clinician, for assessment, intervention, and/or consultation,” is a quickly growing aspect of practicing audiology. However, only 12% of audiologists are involved in providing services via telepractice (REDA International, Inc., 2002). Lack of knowledge regarding telepractice has been cited as one of the reasons many audiologists do not use telepractice to provide audiology services. This study surveyed audiology doctoral students regarding their opinions about the use of telepractice both before and after their opportunity to provide services via telepractice sessions. The authors expected that by providing students the opportunity to have hands-on training in telepractice with supervision, they would be more open to using telepractice after becoming licensed audiologists. Overall, the data indicates benefits of exposing students to telepractice while they are in graduate school.

University Dropout

2008 ◽  
Vol 29 (3) ◽  
pp. 134-147 ◽  
Author(s):  
Manuel C. Voelkle ◽  
Nicolas Sander
Keyword(s):  
Grade Point Average ◽  
Structural Equation ◽  
Unobserved Heterogeneity ◽  
School Grade ◽  
Grade Point ◽  
High School Grade ◽  
Hands On ◽  
University Dropout ◽  
The Individual ◽  
A Prospective Study

University dropout is a politically and economically important factor. While a number of studies address this issue cross-sectionally by analyzing different cohorts, or retrospectively via questionnaires, few of them are truly longitudinal and focus on the individual as the unit of interest. In contrast to these studies, an individual differences perspective is adopted in the present paper. For this purpose, a hands-on introduction to a recently proposed structural equation (SEM) approach to discrete-time survival analysis is provided ( Muthén & Masyn, 2005 ). In a next step, a prospective study with N = 1096 students, observed across four semesters, is introduced. As expected, average university grade proved to be an important predictor of future dropout, while high-school grade-point average (GPA) yielded no incremental predictive validity but was completely mediated by university grade. Accounting for unobserved heterogeneity, three latent classes could be identified with differential predictor-criterion relations, suggesting the need to pay closer attention to the composition of the student population.

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