User:Lauren mun/sandbox

Article evaluation

Electron Impact

[| Electron Impact]

Overall, this article was okay. It was not the best article I have read on Wiki, but it was certainly not the worst. It was a Wiki Education Foundation supported article and was rated for both Physics and Mass Spectrometry WIkiprojects. Overall, It read well in some passages and not others. The passages explaining background, history, and the basics of the ionization process were well-written and concise. They didn’t seem biased or truncated. The applications section of the article did seem a bit jumbled though. No section felt biased, but it seemed a little lopsided and truncated. For example, the section on GC-MS had some weird examples and the one sentence on Vacuum Manifold felt forced. That section in particle seemed very out of place needing to be either expanding or condensed into another paragraph. In general, the applications section seemed a bit off and could use some expansion. The sources used were acceptable and all seemed to work, and the talk page seems dead. Overall, the article could use some editing and refining.

Text I posted on the article’s talk page: Why was the forensics application section only limited to detection of medicinal drugs, and highly specific ones at that? Why not discuss current advances and regular applications of EI in the forensics field? Lauren mun (talk) 23:19, 24 February 2018 (UTC)

Add to an article

-	From section on vacuum manifold sample introduction

This method is useful with highly volatile samples that may not be compatible with other sample introduction methods.

[1] Dass, Chhabil (2007). Desiderio, Dominic; Nibbering, Nico, eds. Fundamentals of Contemporary Mass Spectrometry (1 ed.). Hoboken: John Wiley & Sons, Inc. p. 19.

Initial planning for my article

(Rough Ideas)

EMULSION PCR -	Be careful of non-microdroplet PCR Polymerase Chain Reaction (PCR) has been a vital tool in genomics and biological endeavors, in general, for years. Scaling down PCR to the microscale has allowed single-molecule PCR (Nakano).

IVTT -	Describe -	What is it used for -	Microscale -	Current state /application

DIRECTED EVOLUTION -	Not sure if going to use, focus on high throughput directed evolution by introducing large numbers of mutations simultaneously

Further thought and some sources:

Add and give some examples of work in IVTT. There is not a ton of this specific topic in a concise manner on Wikipedia.

Add and explain microdroplet based PCR techniques since that specific technique is not thoroughly explained on Wikipedia.

Some sources: (just as a starting point)

Griffiths, A. D.; Tawfik, D. S. Miniaturising the Laboratory in Emulsion Droplets. Trends Biotechnol. 2006, 24 (9), 395–402.

Trung, N. B.; Saito, M.; Takabayashi, H.; Viet, P. H.; Tamiya, E.; Takamura, Y. Multi-Chamber PCR Chip with Simple Liquid Introduction Utilizing the Gas Permeability of Polydimethylsiloxane. Sensors Actuators B Chem. 2010, 149 (1), 284–290.

Nakano, M.; Komatsu, J.; Matsuura, S.; Takashima, K.; Katsura, S.; Mizuno, A. Single-Molecule PCR Using Water-in-Oil Emulsion. J. Biotechnol. 2003, 102 (2), 117–124.

Initial drafting of my article

DIRECTED EVOLUTION TECHNIQUES (IVTT, PCR, ETC.)

IVTT

In Vitro compartmentalization (IVC) has allowed for the miniaturization of large-scale techniques that can now be done on the droplet scale. Included in these techniques for directed evolution exploration is in vitro transcription and translation (IVTT). Using droplet techniques allows for high throughput analysis with many different selection pressures in directed evolution. By using a microdroplet technology, fusion of droplets allows for selective introduction of molecules used to influence gene expression (Luke).

PCR

(Link to droplet PCR article)

Polymerase Chain Reaction (PCR) has been a vital tool in genomics and biological endeavors since its inception. Scaling down PCR to the microscale has allowed single-molecule PCR. Using water-in-oil droplets, droplet PCR operates by assembling droplet ingredients pre-formation, forming droplets, thermocycling, and reading results via fluorescence. This technique is capable of running an excess of 2 million PCR reactions and detection of 100,000-fold extra wild-type alleles to mutant alleles. Droplet-based PCR allows more small-scale, high throughput PCR including error-prone PCR. Error-prone PCR allows for a large number of mutations to be introduced in individual reaction drops. This technique is useful in building a DNA library (citation*).

On-chip PCR allows for similar amplification along with minimized reagents and time delays. Traditional PCR uses more reagents and takes longer. This technique has been utilized with PDMS to minimize evaporation of regents while still granting the same throughput Multiplexed, microdroplet PCR has been developed that allows for screening of large numbers of target sequences in bacterial identification (Xu). On-chip PCR allows for excess of 15 x 15 multiplexing. This multiplexing was made possible with immobilized primers for amplification in the chip wells.


 * cannot find the article. Will continue searching for source that I lost.

Article I emailed for peer review

DIRECTED EVOLUTION TECHNIQUES (IVTT, PCR, ETC.)

IVTT

In Vitro compartmentalization (IVC) has allowed for the miniaturization of large-scale techniques that can now be done on the droplet scale. Included in these techniques for directed evolution exploration is in vitro transcription and translation (IVTT)1. Using droplet techniques allows for high throughput analysis with many different selection pressures in directed evolution. By using a microdroplet technology, fusion of droplets allows for selective introduction of molecules used to influence gene expression2 (Luke). IVTT has been used to further explore ribosomal interactions with RNA polymerase as well. Previous explorations of this interactions had to be done in vivo. The importance of the interactions of transcriptional and translational activities has been explored in recent years. Recruitment order can play a part in gene expression3 (Svetlov). Modern techniques allow for a completely isolated, in vitro, analysis. Purified mRNA, RNA polymerase, amino acids, and all necessary transcription and translation factors are contained in a single system. On factor in in vitro analysis is the ease of switching between different couplings of transcription and translation allowing for assessment of ribosomal and polymerase interactions in replication forks4 (Castro-Roa).

PCR

(Link to droplet PCR article)

Polymerase Chain Reaction (PCR) has been a vital tool in genomics and biological endeavors since its inception. Scaling down PCR to the microscale has allowed single-molecule PCR5 (Nakano). Using water-in-oil droplets, droplet PCR operates by assembling droplet ingredients pre-formation, forming droplets, thermocycling, and reading results via fluorescence. This technique is capable of running an excess of 2 million PCR reactions and detection of 100,000-fold extra wild-type alleles to mutant alleles6. Droplet-based PCR allows more small-scale, high throughput PCR including error-prone PCR. Error-prone PCR allows for a large number of mutations to be introduced in individual reaction drops. This technique is useful in building a DNA library (citation*).

On-chip PCR allows for similar amplification along with minimized reagents and time delays. Traditional PCR uses more reagents and takes longer. This technique has been utilized with PDMS to minimize evaporation of regents while still granting the same throughput7 (Trung). Multiplexed, microdroplet PCR has been developed that allows for screening of large numbers of target sequences in bacterial identification8 (Xu). On-chip PCR allows for excess of 15 x 15 multiplexing. This multiplexing was made possible with immobilized primers for amplification in the chip wells9 (Jung). Recent droplet PCR techniques allow for higher accuracy and amplification of small copy number examples than tradition qPCR. The higher accuracy was due to surfactant-doped PDMS as well as a sandwiched glass-PDMS-glass device design. These device properties allowed for more streamlined priming and less water evaporation during PCR cycling10.

* cannot find the article. Will continue searching for source that I lost.

Citations

(1)      Griffiths, A. D.; Tawfik, D. S. Miniaturising the Laboratory in Emulsion Droplets. Trends Biotechnol. 2006, 24 (9), 395–402.

(2)      Luke, C. J. Serpin Production Using Rapid in Vitro Transcription/translation Systems. Methods 2004, 32 (2), 191–198.

(3)      Svetlov, V.; Nudler, E. Unfolding the Bridge between Transcription and Translation. Cell 2012, 150 (2), 243–245.

(4)      Castro-Roa, D.; Zenkin, N. Methodology for the Analysis of Transcription and Translation in Transcription-Coupled-to-Translation Systems in Vitro. Methods 2015, 86, 51–59.

(5)      Nakano, M.; Komatsu, J.; Matsuura, S.; Takashima, K.; Katsura, S.; Mizuno, A. Single-Molecule PCR Using Water-in-Oil Emulsion. J. Biotechnol. 2003, 102 (2), 117–124.

(6)      Hindson, B. J.; Ness, K. D.; Masquelier, D. A.; Belgrader, P.; Heredia, N. J.; Makarewicz, A. J.; Bright, I. J.; Lucero, M. Y.; Hiddessen, A. L.; Legler, T. C.; et al. High-Throughput Droplet Digital PCR System for Absolute Quantitation of DNA Copy Number. ''Anal. Chem. 2011, 83'' (22), 8604–8610.

(7)      Trung, N. B.; Saito, M.; Takabayashi, H.; Viet, P. H.; Tamiya, E.; Takamura, Y. Multi-Chamber PCR Chip with Simple Liquid Introduction Utilizing the Gas Permeability of Polydimethylsiloxane. Sensors Actuators B Chem. 2010, 149 (1), 284–290.

(8)      Xu, Y.; Yan, H.; Zhang, Y.; Jiang, K.; Lu, Y.; Ren, Y.; Wang, H.; Wang, S.; Xing, W. A Fully Sealed Plastic Chip for Multiplex PCR and Its Application in Bacteria Identification. Lab Chip 2015.

(9)      Jung, S.; Kim, B. K.; Lee, S.; Yoon, S.; Im, H.-I.; Kim, S. K. Multiplexed on-Chip Real-Time PCR Using Hydrogel Spot Array for microRNA Profiling of Minimal Tissue Samples. Sensors Actuators B Chem. 2018, 262, 118–124.

(10)    Fu, Y.; Zhou, H.; Jia, C.; Jing, F.; Jin, Q.; Zhao, J.; Li, G. A Microfluidic Chip Based on Surfactant-Doped Polydimethylsiloxane (PDMS) in a Sandwich Configuration for Low-Cost and Robust Digital PCR. Sensors Actuators B Chem. 2017, 245, 414–422.

[Link to articles for transcription, translation, PCR]

Revised article after peer review

In Vitro Transcription and Translation
In vitro compartmentalization (IVC) enables the miniaturization of large-scale techniques that can now be done on the droplet scale. One technique for directed evolution exploration is in vitro transcription and translation (IVTT). Droplet techniques enable high throughput analysis with many different selection pressures in directed evolution experimentation. By using microdroplet technology, fusion of droplets allows for selective introduction of molecules used to influence gene expression. IVTT in microdroplets is preferred when overexpression of a desired protein would be toxic to a host cell minimizing the utility of the transcription and translation mechanisms.

Applications in Molecular Genetics
IVTT has been used to further explore ribosomal interactions with RNA polymerase while previous explorations of these interactions had to be done in vivo before the isolation of non-cellular transcription and translation mechanisms; since the machinery responsible for transcription and translation had not yet been isolated from the rest of the cellular contents. The importance of the interactions of transcriptional and translational activities has been explored in recent years using in vitro techniques finding that recruitment order of replication machinery can play a part in gene expression. Modern techniques allow for a completely isolated, in vitro, analysis. Purified mRNA, RNA polymerase, amino acids, and all necessary transcription and translation factors are contained in a single system. One factor in in vitro analysis is the ease of switching between different couplings of transcription and translation allowing for assessment of ribosomal and polymerase interactions in replication forks.

Microdroplet Emulsion Polymerase Chain Reaction
Polymerase chain reaction

Polymerase chain reaction (PCR) has been a vital tool in genomics and biological endeavors since its inception as it has greatly sped up production of DNA samples for a wide range of applications. Scaling down PCR to the microscale has allowed single-molecule PCR on a chip device. Using water-in-oil emulsions, droplet PCR operates by assembling ingredients, forming droplets, thermocycling, and reading results via fluorescence or other methods of detection. This technique is capable of running an excess of 2 million PCR reactions and detecting of 100,000-fold extra wild-type alleles to mutant alleles. Droplet-based PCR enables smaller-scale, high throughput PCR including error-prone PCR. Error-prone PCR allows for the introduction of a large number of mutations to individual reaction droplets. This technique is useful in building a DNA library.

On-chip PCR allows for similar amplification along with minimized reagents and time delays. This technique has been utilized with polymethylsiloxane (PDMS) to minimize evaporation of reagents while still granting the same throughput. Multiplexed, microdroplet PCR has been developed that allows for screening of large numbers of target sequences including such applications as bacterial identification. On-chip PCR allows for an excess of 15 x 15 multiplexing meaning that multiple target DNA sequences could be run on the same device at the same time. This multiplexing was made possible with immobilized DNA primer fragments placed in the base of the individual wells of the chips. Recent droplet PCR techniques allow for higher accuracy and amplification of small copy number examples in comparison to traditional quantitative PCR experiments. The higher accuracy was due to surfactant-doped PDMS as well as a sandwiched glass-PDMS-glass device design. These device properties allowed for more streamlined priming and less water evaporation during PCR cycling. Reflective essay

Write a "reflective essay" (<1.5 pages) on your Wikipedia contributions. Your concise summary here will also help us assign points for the "Final article" and "Peer review" portions of your grade. Please create a section in your sandbox entitled "Reflective essay," and answer the following questions (numbering them as below):

1. What article did you work on? Was this a new article or    an existing article?

I worked on a section for the Droplet Based Microfluidics article on the topic of directed evolution techniques including IVTT and PCR. This was an existing article, but it did not have much on biological applications.

2. Summarize your main contributions in 3-4 sentences or bullet points.

·     In vitro transcription and translation basics as used with droplet based microfluidics

·     Examples of applications of IVTT in molecular genetics including work in transcription and translation machinery recruitment as an example

·     Explanation of microdroplet based PCR in W/O emulsions

·     Extension of on-chip PCR technology including multiplexing abilities

3. How did you respond to suggestions from peer reviewers? Please list specific changes in 3-5 sentences or bullet points. Also    indicate if you used the Wikipedia content expert or received feedback     from other Wikipedians outside the course.

I heavily relied on peer reviews from my classmates. They provided excellent feedback on both technical aspects and structure. I did not use any outside feedback.

·     Split my IVTT section up into two paragraphs and added more details to IVTT

·     Clarified technical language throughout and expanded some technical background

·     Corrected a couple of missed citations in both sections of my work

·     Vastly improved grammar and sentence structure that was missed on my proof reading

4. Reflect on the following questions in a short    paragraph: Was this assignment valuable to your learning (of course     material, research/literature review skills, ability to critically     evaluate peers, etc.) - why or why not? Do you think your article will be    valuable to Wikipedia readers? How could this assignment be improved in    the future? [You will not lose points for negative comments; please be honest    in your critiques of this assignment to improve the course for future     years.]

This article was incredibly valuable to my graduate education as I have not had to write an article like this before. I have never had to concisely explain scientific ideas in such a manner, so I had to really change how I approached an article before I was able to write my Wikipedia content. This assignment also helped me integrate peer review material in a more helpful manner. Since I did not get a chance to evaluate my own peers, I took some extra time while reviewing my corrections to see how other people reviewed my work. I think this is a unique assignment that forced a new, but necessary, writing style. I am not entirely sure if my additions to Wikipedia will be helpful to a large audience, but they will certainly be helpful to the select group who needs concise information on droplet based microfluidics and a lead on further sources.

This assignment was, in the end, very helpful. Throughout the quarter, it was frustrating to keep up with the deadlines as it was separated from class material more than is usually comfortable for me. I don’t love how the Wikipedia timeline interface is set up, so it took me a while to get used to it. My biggest struggle with this assignment was the lack of integration to the course structure. It seemed to be parallel and nonintegrated for most of the quarter. This assignment on top of the snapshots, review, and design projects was a lot to keep track of. Had everything been integrated on one platform it would have been easier to track logistically. The other thing that seemed sort of tacked on at the end was the article editing portion. That assignment was only mentioned briefly in the beginning and then nothing was said again until right before the due date of the wiki articles. It has been rather frustrating as a student to have all of these bits and pieces in different calendars. It feels like two separate classes worth of assignments that haven’t been fully integrated. That is more of a logistical complaint than a critic of the work itself. Now that I have finished my article additions, I am glad we had this assignment, and I feel like it was an assignment that contributed to work outside of the classroom. It was much preferred to normal exams.