#cDNA synthesis

[saiboubaiyou]

合成とPCR

Preparation Reagents

・ISOGENⅡ 　　　　　　　　　　　　　

・2mL tube

・RNA free water 　　　　　

・1.5mL tube

・p-Bromoanisole 　　　　　　　　　　　　

・Tube stand (blue letters)

・Isopropanol  　　　　　　　　　　　　　

・10、100、1000μL tip

・75% ethanol 　　　　　　　　　　　　　

・Micropipettes 5 kinds (6 kinds)

To be done before the experiment

UV should be applied for at least 2 hours.

(If you start in the morning, you can start the night before.)

Preparation of necessary reagents

・Ethanol: The prepared ethanol is 99.5% ethanol. Dilution is necessary.

Method: Dilute 35 mL of ethanol and 15 mL of RNase free water in a 50-mL tube (white lid).

Dilute with 35 mL of ethanol and 15 mL of RNase free water.

・Isopropanol: Dispense into a 50-mL tube.

・Preparation of RNase free water

tRNA measurement

① Dispense 1 mL of ISOGENⅡ into a 2 mL tube.

② Aspirate the medium, put ① into Dish, peel off the cell, and return it to the 2 mL tube.

(Can be stored at -80°C)

③ Replace the aspirator pasteur and gallon bottle.

④ Add 400 μL (0.4 mL) of RNAse free water into the tube (2).

⑤ Shake ④ vigorously by hand for 15 seconds. Then, leave at room temperature for 5 to 15 minutes.

⑥ Centrifuge at 4°C, Max speed, for 15 minutes.

(Put 3.5 μL of p-bromoanisole in another 1.5-mL tube.)

⑦ Take 700 μL of the supernatant and put it into the tube containing p-bromoanisole. Then, shake vigorously for 15 seconds and leave in a clean bench for 5 minutes.

⑧ Centrifuge at 4°C, Max speed, for 10 minutes.

⑨ Put 500 μL of isopropanol in another 1.5-mL tube.

(Write sample name, date, name, and tRNA on the lid.)

⑩ Take 500 μL of the supernatant of ⑧, add it to ⑨, mix by inversion, and leave for 10 minutes.

⑪ Centrifuge at 4°C, max speed, for 10 minutes.

⑫ Aspirate the supernatant.

⑬ Add 0.5mL 75% ethanol, centrifuge at 4°C, max speed, for 5 minutes.

⑭ Aspirate the supernatant.

⑮ Perform steps 13 and 14 again.

⑯ Centrifuge (1 min) to increase the purity of the pellet. (Take the excess supernatant with a 1 µL tip)

⑰ Cover the tube with a prowipe with the lid open and leave it in the clean bench for 10 minutes.

(Turn off the fan of the clean bench.) (Drying is complete when the pellet turns from white to clear)

Dissolve (pipetted) in ⑱ 8 µL RNAse free water

(The RNA can be stored at -20°C, but since RNA degrades quickly, it is recommended that the cDNA be prepared within one day.)

⑲ Flushing

⑳ 20 μL of RNA free water in a 1.5 mL tube for tRNA measurement*.

*tRNA measurement method

① Open a dedicated PC

② Click the Nano Drop 2000 icon.

③Select "No" in the Question window and "OK" in the next window.

④ Confirm that the lid of the instrument is closed, and select "Nucleic Acid".

⑤ Remove all ✔︎ on the left side of the screen.

⑥ Select "RNA" for Target on the right side of the screen.

⑦Set the unit to ng/μL. (8) Open the lid of the analyzer and select "Nucleic Acid".

⑧Open the lid of the analyzer, pour 1 μL of RNA free water into the measurement site, wash the site twice, and click "blank" in the upper left corner of the screen.

⑨Add 1 μL of sample and click "Measurement" in the upper left corner.

→Input the values of concentration, 260/280 and 260/230 into your Excel.

　Measure each sample twice and take the average.

⑩When finished, rinse the sample twice with water and turn off the PC.

cDNA Synthesis Method

Preparation

・Set up the incubator in advance.

・Dilute Random Primer 10-fold with TE Buffer.

・Dilute dNTPs 2.5x in TE Buffer.

Methods.

① Add 5Buffer, 10mM dNTPs, Revertra Ase, and RNAse Inhibitior in this order to a 1.5mL tube.

② Add 1 μL of Random Primer to a new 1.5 mL tube.

③ Add the sample and RNAse free water to (2), based on the input for the tRNA concentration measurement.

④ Incubate at 65℃ for 5 minutes. Then, open the lid, immediately hold the tube and place the tube on ice.

⑤ Flush the sample and put 6.75 μL of mix solution.

⑥ Annealing 30℃, 10 min.

⑦ Enzyme reaction 42℃, 60 min.

⑧ Transformation 99℃, 5 min.

⑨ Put the sample on ice, flush, and transfer to the freezer.

Mix solution for cDNA synthesis (make a little extra)

5×Buffer = 4

10 mM dNTPs = 2

ReverTra Ace = 0.25

RNAse Inhibitor = 0.5

total = 20

Unit: μL

RT-qPCR method

① Put 30 μL of TE buffer into a tube.

② Add 20 μL of primer stock solution.

③ Vortex the tubes, then add water, primers (Forward, Reverse), and Luna in this order.

④ Mark the 8 tubes and put in the mix solution. (14.7 µL) ⑤ Put in the sample.

⑤ Add sample (0.3 µL). (0.3 µL) ⑥ Put the sample into the PCR machine.

⑥ Put the sample into the PCR device**.

Mix solution

Luna = 7.5

Forward = 0.18

Reverse = 0.18

RNA free水 = 6.84

RT-qPCR instrument

① Click on "New experiment

② Click on "Comparative quantitation

③ Set the tube location.

④ Enter ✔︎ in "SYBR" at "Add Dyes". (5) Adjust the temperature in "Thermal profile".

⑤ Adjust the temperature in "Thermal profile" (6) Click "+" to set the temperature.

⑥ Click "+" and press "melt" to see the melt curve (accuracy index).

⑦ Click "Run experiment" and click "Save? →Click "OK".

⑧ Select "hattori", choose your file, and put in the title, date, etc.

⑨ Check if the place you want to measure is surrounded by a bold black line and if "SYBR" is included.

⑩ Click "save" to start measurement.

[transgenderer]

this article is really cool, ive never seen such a radically biological study of memory! also, check out the properties of their model organism:

After an extensive search for a suitable experimental animal, I settled on the giant marine snail Aplysia (Fig. 1A) because it offers three important advantages: Its nervous system is made up of a small number of nerve cells; many of these are gigantic; and (as became evident to me later) many are uniquely identifiable (3, 4). Whereas the mammalian brain has a trillion central nerve cells, Aplysia has only 20,000, and the simplest behaviors that can be modified by learning may directly involve less than 100 central nerve cells. In addition to being few in numbers, these cells are the largest nerve cells in the animal kingdom, reaching up to 1000 mmin diameter, large enough to be seen with the naked eye. One can record from these large cells for many hours without any difficulty, and the same cell can be returned to and recorded from over a period of days. The cells can easily be dissected out for biochemical studies, so that from a single cell one can obtain sufficient mRNA to make a cDNA library. Finally, these identified cells can readily be injected with labeled compounds, antibodies, or genetic constructs, procedures which opened up the molecular study of signal transduction within individual nerve cells.

also some remarkable results about determinism in neural development

Kupfermann, Castellucci, Carew, Hawkins, John Byrne, and I worked out significant components of the neural circuit gill-withdrawal reflex (Fig. 2). The circuit is located in the abdominal ganglion and has 24 central mechanoreceptor sensory neurons that innervate the siphon skin and make direct monosynaptic connections with 6 gill motor cells (Fig. 2C) (12–14). The sensory neurons also made indirect connections with the motor cells through small groups of excitatory and inhibitory interneurons (15, 16). In addition to being identifiable, individual cells also have surprisingly large effects on behavior (Fig. 2B) (4, 14, 17). As we examined the neural circuit of this reflex, we were struck by its invariance. In every animal we examined, each cell connected only to certain target cells and not to others (Fig. 2C). This also was true in the neural circuitry of other behaviors in Aplysia including inking, control of the circulation, and locomotion (4, 18)

anyway the big takeaways here are

you can get long-term learning without changing the connections between neurons if you change the *strength* of the connections between neurons (and you can directly measure these changes)

short term memory forms even when protein synthesis is inhibited, but long term doesnt, and they were actually able to piece out which protein cascades (like, you go and transcribe one protein from the DNA, then this starts a process that transcribes other proteins, etc) are involved!

another detail, about how you can have synapse-specific changes when stuff is going on in the nucleus, which shares a huge number of synapses:

The finding of a transcriptional cascade explained why long-term memory requires new protein synthesis immediately after training, but it posed a new cell-biological problem. A single neuron makes hundreds of contacts on many different target cells. Short-term synaptic changes are synapse-specific. Since longlasting synaptic changes require transcription and thus the nucleus, is long-term memory storage a cell-wide process, or are there cellbiological mechanisms that maintain the synapse specificity of long-term facilitation? To examine these questions, Kelsey Martin cultured one Aplysia sensory cell with a bifurcating axon with two motor neurons, forming two widely separated synapses (Fig. 4A). In this culture system, a single puff of serotonin applied to one synapse produces transient facilitation at that synapse only, as expected (50, 51). Five puffs of serotonin applied to one branch produces long-lasting facilitation (72 hours), also restricted to the stimulated synapse (Fig. 4B). This long-lasting synapse-specific facilitation requires CREB and also leads to structural changes. Thus, despite recruitment of nuclear processes, long-term changes in synaptic function and structure are confined only to those synapses stimulated by serotonin. How does this come about? Martin, Andrea Casadio, Bailey, and I found that five puffs of serotonin send a signal to the nucleus to activate CREB-1, which then appears to send proteins to all terminals; however, only those terminals that have been marked by serotonin can use these proteins productively for synaptic growth. Indeed, one puff of serotonin to the previously unstimulated synapse is sufficient to mark that synapse so that it can capture a reduced form of the long-term facilitation induced at the other site by five puffs of serotonin (Fig. 4B)

[buggbuzz]

You should be allowed to kill during the exam. Just 1 person

this is actually really funny cause it's reminding me of how the professor said she added a couple problems just because my friend kept asking her specific questions

and everyone turned to look at him and he was just like "...everyone gets one kick 😞" LMAOOOO

anyways the exam wasn't objectively terrible, it was lowkey my fault for letting my stardew fixation get in the way, but wow talk about a thorough ass test! the short response wanted a full flow chart of the process of RT-PCR (with enzymes and temperatures) and the long response wanted a full explanation of how i would synthesize a hormone (EPO) excreted from the human kidney that required glycosylation with thorough details on how each step and technique worked and why i picked them 😭😭😭

i actually got a perfect score on the short response and almost a perfect score on the long response but i was so unsure of how exactly cloning worked and didnt wanna say something false that i glossed over that which cost me 8/10 points for that part💔💔

since i really wanna talk abt this tho cause im lowkey proud of what i came up with:

since a human hormone is probably produced by a pretty large gene i'd want something bigger than a regular plasmid or bacteriophage. since i need proteins to be produced, a cosmid or ti vector are also off the table (because the viral capsid and plant cell wall would greatly interfere with extraction), and since i need a post-translational modification (glycosylation) performed i need a eukaryotic vector, eliminating bacterial artificial chromosomes (BACs), which leaves the best vector as a yeast artificial chromosome (YAC).

then, because i figured if any gene was gonna have alternative gene splicing it was gonna be the one for some fancy human hormone that regulates blood synthesis, i wanted to extract an mRNA transcript (if i'd just said i would isolate it from a pig kidney or smth i would've gotten more points😭😭😭) that i'd then synthesize cDNA for through RT PCR to eliminate any introns.

since that was like 90% of what i could guess was necessary without more information about EPO or yeast transformation, i kinda just picked random extraction, chromatography, and verification techniques for the rest of it. i said i'd assume that the yeast would excrete the protein like the kidney cells would, meaning i'd just extract the buffer, use some salts to dehydrate the buffer so the hydrophobic proteins would clump together, centrifuge it to make them collect into a pellet, use size-exclusionary chromatography to isolate the presumably-large EPOs from the other proteins, then use X ray crystallography to verify the protein extracted was indeed EPO. i really wanted to mention western blotting but i wasn't very sure of how i'd describe it beyond the double antibody thing (which i LOVE) so i didnt wanna risk it lol.

[articlepublication]

SPRI Bead-Based Purification: Optimizing PCR Cleanup for Superior NGS Results

In molecular biology, precision and purity are everything. Whether you’re prepping for next-generation sequencing (NGS), cloning, or qPCR, the quality of your DNA directly influences your results. PCR cleanup is one of those essential steps that can make or break your downstream applications. And when it comes to PCR cleanup, Solid Phase Reversible Immobilization (SPRI) bead-based purification is the method of choice for researchers worldwide.

SPRI bead-based methods combine efficiency, scalability, and versatility in one simple workflow. They’ve become indispensable for high-throughput labs, small academic teams, and commercial genomic facilities alike. In this article, we’ll explore how SPRI beads work, best practices for optimizing PCR cleanup, and how cost-effective solutions like MagBio Genomics’ HighPrep™ PCR Clean-Up Kit are transforming the purification landscape.

What is SPRI Technology?

SPRI stands for Solid Phase Reversible Immobilization. This method uses paramagnetic beads that bind nucleic acids in the presence of polyethylene glycol (PEG) and salt. The DNA binds to the beads, while unwanted components like salts, primers, enzymes, and nucleotides remain in the solution.

Key benefits include:

Size-selective binding

High recovery rates for DNA fragments ranging from 100 bp to >10 kb

Simple workflows that replace spin-column protocols

Automation-friendly—ideal for 96- and 384-well plates

The Role of PCR Cleanup in NGS Workflows

NGS library preparation involves several steps that require precise DNA handling. After amplification, it’s critical to remove unwanted materials such as leftover primers, free nucleotides, and enzymes. These contaminants can:

Inhibit adapter ligation

Interfere with quantification and normalization

Reduce overall sequencing quality

SPRI beads simplify this process. Add the beads, bind the DNA, wash away the contaminants, and elute clean DNA that’s ready for the next step.

Key Advantages of SPRI-Based PCR Cleanup

1. Efficiency

SPRI workflows reduce the number of steps and eliminate centrifugation. This makes them ideal for high-throughput pipelines and minimizes user error.

2. Scalability

From a handful of samples to thousands, SPRI bead protocols are easily scaled and automation-compatible.

3. Customizability

You can fine-tune your purification with bead-to-sample ratios:

1.8x for total DNA recovery

0.6x to exclude smaller fragments (<300 bp)

0.8x to retain larger DNA for long-read workflows

How to Optimize SPRI Bead-Based PCR Cleanup

Choose the Right Bead-to-Sample Ratio

This is perhaps the most important variable. Use:

1.8x for complete fragment retention

0.8x to remove primer dimers and small artifacts

Avoid Overdrying the Beads

After ethanol washing, beads should be air-dried until no visible liquid remains. Overdrying can make it difficult to elute DNA and decrease recovery.

Use Fresh Ethanol

Old or cold ethanol can decrease the wash efficiency and lead to poor DNA purity.

Elute in the Right Volume

Using 20–50 μL of elution buffer ensures efficient recovery while keeping the DNA concentration high enough for downstream use.

Common Applications in Molecular Biology

SPRI beads aren’t just for NGS. Their versatility spans across:

PCR cleanup: Removing enzymes, primers, dNTPs

Cloning: Preparing inserts for ligation

Sanger sequencing: Enhancing read clarity by removing noise

qPCR: Ensuring template purity for quantification

cDNA synthesis and RNA-seq: Purifying double-stranded cDNA

HighPrep™ PCR Clean-Up Kit: A Reliable SPRI-Based Solution

MagBio Genomics’ HighPrep™ PCR Clean-Up Kit is designed to deliver all the benefits of SPRI bead-based purification at a more accessible cost.

Key Features:

Compatible with all major NGS platforms

Comparable or better recovery than AMPure XP

Validated for manual and automated protocols

Stable shelf life and consistent batch quality

HighPrep™ beads use the same SPRI chemistry as traditional solutions but come at a fraction of the cost, making them perfect for cost-sensitive labs without sacrificing performance.

Automation and High-Throughput Compatibility

As research scales up, automation becomes essential. HighPrep™ PCR Clean-Up is compatible with liquid-handling platforms like:

Thermo Fisher KingFisher Flex

Hamilton Microlab STAR

Tecan Fluent and Evo

These integrations allow genomic core facilities to streamline operations and reduce sample-to-sample variation.

Testimonials and Case Studies

“Switching to HighPrep™ not only cut our cleanup costs in half, but also gave us flexibility with automation. We saw no drop in yield or data quality.” — Senior Scientist, Biotech Startup

“We routinely process 10,000 samples a month, and HighPrep™ has been instrumental in keeping our workflow smooth and cost-effective.” — Genomics Core Facility, UK

Sustainability and Supply Chain Reliability

Unlike spin columns, SPRI beads reduce plastic usage. HighPrep™ PCR Clean-Up is also offered in bulk formats to minimize packaging waste. And unlike some competitors, MagBio ensures consistent availability with a stable supply chain.

Cost Comparison

When to Use SPRI-Based Purification

SPRI bead cleanup is ideal when you need:

Fast and reproducible DNA purification

High throughput compatibility

Flexible size selection for fragment trimming or retention

Reliable performance for sensitive applications like NGS

Final Thoughts

SPRI bead-based purification has changed the game for PCR cleanup and beyond. The simplicity, accuracy, and scalability of this approach make it indispensable in modern molecular biology. With cost-effective and high-performing kits like HighPrep™ PCR Clean-Up from MagBio Genomics, it’s easier than ever to streamline your workflow, reduce costs, and get consistent, high-quality results.

Upgrade Your PCR Cleanup Workflow

Explore the power of SPRI-based purification with the HighPrep™ PCR Clean-Up Kit. It’s the perfect solution for efficient, cost-effective DNA purification in any lab setting.

[global-research-report]

Exploring the Latin America Gene Expression Market

The Latin America gene expression market size is expected to reach USD 578.3 million by 2027, according to a new report by Grand View Research, Inc. The market is expected to expand at a CAGR of 5.6% from 2020 to 2027. High investment in alternative approaches for drug discovery and an increased number of projects involving genetic studies are expected to drive demand.

The region is characterized by fragmented healthcare and a high rate of poverty as well as disparities to access to standard care for cancer and other complex disorders. In this region, consumers from public settings are unable to afford targeted therapies. The number of facilities providing genomic platforms is limited in the region. These factors have resulted in the low revenue share of the Latin America market in the global market for gene expression.

However, researchers and healthcare providers are pursuing training for the use of genomic platforms and constantly participating in genomic projects. Currently, a medical practice based on precision medicine is still a challenge and limited to private settings in Latin America. A rise in the number of research studies pertaining to gene expression, genomic medicine, and personalized medicine in the LATAM region is expected to drive the market.

The Brazilian Initiative on Precision Medicine (BIPMed) is the first Latin American and Brazilian beacon, which is a genetic mutation sharing platform established by the Global Alliance for Genomics and Health (GA4GH). BIPMed encourages propagation and guidance in areas of computational ecology, personalized medicine, molecular hereditary, and others. This initiative plays an important role in the development of precision medicine in Latin America, thereby enabling significant advancements in healthcare and research in this region.

Latin America Gene Expression Market Report Highlights

cDNA synthesis and conversion accounted for the largest revenue share in 2019. The introduction of systems that provide direct library synthesis eliminating the need for prior steps is expected to increase the revenue share of the segment

Kits and reagents generated the largest product revenue in 2019 owing to the commercial availability of a broad range of kits and reagents to support biological discoveries and advancements

Moreover, the introduction of enhanced kits and reagents that support better workflow, also contributes to segment growth

Availability of products with higher efficiency, better adoption to specific research need, and decline in prices of associated techniques is projected to drive growth in the coming years

High-Plex capacity is expected to witness the fastest CAGR from 2020 to 2027. The segment also accounted for the largest revenue share owing to high adoption in clinical and drug discovery uses

Increased demand for advanced diagnostic solutions is expected to fuel the growth of the clinical diagnostics segment

RNA expression dominated the technique segment. Technological advances in RNA expression tools and techniques coupled with the availability of a range of products to meet different research needs are anticipated to increase the adoption of these techniques for gene expression analysis

Brazil accounted for the largest revenue generation in 2019 owing to continuous developments in genomicand healthcare-related research in the country

Competition in the market is high due to the presence of a substantial number of well-established companies and small- to mid-sized entities

Key players are manufacturing and commercializing products to cater to various requirements in gene expression. To maintain a significant share in the market, players have implemented several business strategies including the expansion of distribution network, product portfolio, and increase in manufacturing capacity through mergers and collaborations

Latin America Gene Expression Market Segmentation

Grand View Research has segmented the Latin America gene expression market on the basis of process, product, capacity, application, technique, and region:

Latin America Gene Expression Process Outlook (Revenue, USD Million, 2016 - 2027)

Sample Collection

Purification

cDNA Synthesis & Conversion

PCR Analysis

Data Analysis & Interpretation

Latin America Gene Expression Product Outlook (Revenue, USD Million, 2016 - 2027)

Kits & Reagents

DNA Chips

Others

Latin America Gene Expression Capacity Outlook (Revenue, USD Million, 2016 - 2027)

Lolex 

Latin America Gene Expression Application Outlook (Revenue, USD Million, 2016 - 2027)

Drug Discovery & Development

Clinical Diagnostics

Biotech & Microbiology

Others 

Latin America Gene Expression Technique Outlook (Revenue, USD Million, 2016 - 2027)

RNA Expression

Northern Blotting

DNA Microarrays

PCR Techniques

RNA-Sequencing

Promoter Analysis

In vitro Transcription/Nuclear Run-On Assays

Gel Shift Assays

Chromatin Immunoprecipitation (ChIP)

Protein Expression & Posttranslational Modification Analysis

Western Blotting

2-D Gel Electrophoresis

Immunoassays

Mass Spectrometry

Latin America Gene Expression Regional Outlook (Revenue, USD Million, 2016 - 2027)

Latin America

Brazil

Mexico

Argentina

Colombia

Chile

Peru

Ecuador

Dominican Republic (República Dominicana)

Venezuela

Guatemala

Panama

Costa Rica

Uruguay

Cuba

Order a free sample PDF of the Latin America Gene Expression Market Intelligence Study, published by Grand View Research.

[healthcaremarketanalysis]

Exploring RNA Analysis Methods: Techniques for Comprehensive Understanding of RNA

RNA analysis is a cornerstone of molecular biology, enabling researchers to decode the various functions and regulatory mechanisms of RNA in cellular processes. With growing interest in transcriptomics, RNA analysis methods have evolved to offer more precise, high-throughput, and comprehensive insights into gene expression, alternative splicing, RNA modifications, and more. Here, we explore several RNA analysis methods that have become essential tools in biological and medical research.

Download PDF Brochure

1. RNA Sequencing (RNA-Seq)

RNA sequencing is the gold standard for transcriptome analysis. It allows researchers to examine both coding and non-coding RNA with high resolution. RNA-Seq provides quantitative data on gene expression levels, alternative splicing events, and even RNA-editing phenomena. This method has the advantage of being unbiased, offering a comprehensive snapshot of the entire transcriptome.

Steps Involved:

RNA extraction

cDNA synthesis

Sequencing via next-generation sequencing platforms

Data analysis using bioinformatics tools to map reads to reference genomes and quantify expression

2. Quantitative PCR (qPCR)

Quantitative PCR is a highly sensitive method to measure RNA expression levels. It is often used to validate results from RNA-Seq or microarray studies. By amplifying specific RNA sequences and using fluorescent probes, qPCR provides real-time quantification of RNA molecules, offering highly accurate and reproducible data.

Advantages:

High sensitivity

Quantitative results in real time

Often used for validation of gene expression studies

3. Microarrays

Microarray technology allows the simultaneous analysis of thousands of RNA molecules. Although it has been somewhat replaced by RNA-Seq due to the latter’s higher resolution and broader coverage, microarrays remain popular for focused studies on specific genes or pathways. They are relatively inexpensive and easy to use for researchers looking for rapid gene expression profiling.

Key Applications:

Gene expression profiling

Comparative studies across different samples or conditions

Focused analysis of known RNA sequences

4. Northern Blotting

Northern blotting is a classical technique used to detect specific RNA molecules within a mixture of RNA. While it is less commonly used today, northern blotting remains a reliable tool for detecting the presence and size of RNA molecules. This method is particularly useful for validating the results of RNA-Seq or qPCR.

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Process Overview:

RNA extraction and electrophoresis

Transfer of RNA onto a membrane

Hybridization with labeled probes specific to the RNA of interest

Detection via autoradiography or chemiluminescence

5. Single-Cell RNA Sequencing (scRNA-Seq)

Single-cell RNA sequencing is a cutting-edge technique that enables researchers to study gene expression at the resolution of individual cells. This method has revolutionized the field of transcriptomics by revealing cellular heterogeneity and identifying rare cell types that might be missed by bulk RNA-Seq.

Advantages:

High resolution for detecting cell-to-cell variability

Crucial for understanding complex tissues and diseases like cancer

Insights into cellular differentiation and development

6. RNA Immunoprecipitation (RIP)

RNA immunoprecipitation is used to study RNA-protein interactions. Researchers use specific antibodies to target RNA-binding proteins, isolating the associated RNA molecules. RIP is particularly valuable in studying RNA modifications, such as methylation, and understanding how RNA-protein complexes influence gene expression.

Applications:

Studying RNA modifications (e.g., m6A methylation)

Understanding the role of RNA-binding proteins in disease

Functional annotation of RNA molecules

7. In Situ Hybridization (ISH)

In situ hybridization is a method used to detect specific RNA sequences in fixed tissue sections or cells. This method provides spatial information about RNA localization within tissues, making it invaluable for developmental biology and cancer research.

Benefits:

Visualization of RNA expression patterns in intact tissues

High spatial resolution

Useful in identifying RNA localization in specific cell types

Conclusion

The diversity of RNA analysis methods allows researchers to study the complex roles of RNA in gene regulation, cellular function, and disease. While RNA-Seq remains the most comprehensive approach, each method offers distinct advantages depending on the research question and experimental needs. By combining these methods, scientists can gain a holistic view of RNA biology, paving the way for advancements in precision medicine and therapeutic development.

Whether it's detecting subtle changes in gene expression or unraveling RNA-protein interactions, these RNA analysis techniques continue to enhance our understanding of the molecular underpinnings of life.

Content Source:

[jhavelikes]

Defense-associated reverse transcriptase (DRT) systems perform DNA synthesis to protect bacteria against viral infection, but the identities and functions of their DNA products remain largely unknown. Here we show that DRT2 systems encode an unprecedented immune pathway that involves de novo gene synthesis via rolling circle reverse transcription of a non-coding RNA (ncRNA). Programmed template jumping on the ncRNA generates a concatemeric cDNA, which becomes double-stranded upon viral infection. Remarkably, this DNA product constitutes a protein-coding, nearly endless ORF (neo) gene whose expression leads to potent cell growth arrest, thereby restricting the viral infection. Our work highlights an elegant expansion of genome coding potential through RNA-templated gene creation, and challenges conventional paradigms of genetic information encoded along the one-dimensional axis of genomic DNA.

De novo gene synthesis by an antiviral reverse transcriptase | Science

[leedsomics]

Scalable spatial single-cell transcriptomics and translatomics in 3D thick tissue blocks

Characterizing the transcriptional and translational gene expression patterns at the single-cell level within their three-dimensional (3D) tissue context is essential for revealing how genes shape tissue structure and function in health and disease. However, most existing spatial profiling techniques are limited to 5-20 m thin tissue sections. Here, we developed Deep-STARmap and Deep-RIBOmap, which enable 3D in situ quantification of thousands of gene transcripts and their corresponding translation activities, respectively, within 200-m thick tissue blocks. This is achieved through scalable probe synthesis, hydrogel embedding with efficient probe anchoring, and robust cDNA crosslinking. We first utilized Deep-STARmap in combination with multicolor fluorescent protein imaging for simultaneous molecular cell typing and 3D neuron morphology tracing in the mouse brain. We also demonstrate that 3D spatial profiling facilitates comprehensive and quantitative analysis of tumor-immune interactions in human skin cancer. http://dlvr.it/TBf98L

[rnomics]

Discrete measurements of #RNA polymerase and reverse transcriptase fidelity reveal #evolutionary tuning [Method]

Direct methods for determining the fidelity of DNA polymerases are robust, with relatively little sample manipulation before sequencing. In contrast, methods for measuring RNA polymerase and reverse transcriptase fidelities are complicated by additional preparation steps that introduce ambiguity and error. Here, we describe a sequencing method, termed Roll-Seq, for simultaneously determining the individual fidelities of RNA polymerases and reverse transcriptases (RT) using Pacific Biosciences Single Molecule Real-Time sequencing. By employing reverse transcriptases with high rolling-circle activity, Roll-Seq generates long concatemeric cDNA from a circular RNA template. To discern the origin of a mutation, errors are recorded and determined to occur within a single concatemer (reverse transcriptase error) or all concatemers (RNA polymerase error) over the cDNA strand. We used Roll-Seq to measure the fidelities of T7 RNA polymerases, a Group II intron-encoded RT (Induro), and two LINE RTs (Fasciolopsis buski R2-RT and human LINE-1). Substitution rates for Induro and R2-RT are the same for cDNA and second strand synthesis while LINE-1 has 2.5-fold lower fidelity when performing second strand synthesis. Deletion and insertion rates increase for all RTs during second strand synthesis. In addition, we find that a structured RNA template impacts fidelity for both RNA polymerase and RT. The accuracy and precision of Roll-Seq enable this method to be applied as a complementary analysis to structural and mechanistic characterization of RNA polymerases and reverse transcriptases or as a screening method for RNAP and RT fidelity. http://rnajournal.cshlp.org/cgi/content/short/rna.080002.124v2?rss=1&utm_source=dlvr.it&utm_medium=tumblr

[wellnessweb]

MicroRNA Market Size: Opportunities and Challenges

The MicroRNA Market size was USD 1.57 Billion in 2023 and is expected to Reach USD 4.13 Billion by 2031 and grow at a CAGR of 12.88% over the forecast period of 2024-2031.The microRNA (miRNA) market is experiencing significant growth, driven by the increasing recognition of miRNAs as critical regulators of gene expression and their potential as biomarkers and therapeutic targets.

Advances in miRNA research have opened new avenues for precision medicine, particularly in oncology, where miRNA-based diagnostics and therapies offer promising approaches for early detection and personalized treatment. The market is also benefiting from technological innovations in next-generation sequencing (NGS) and bioinformatics, which enhance miRNA profiling and functional analysis. Additionally, the growing investment in biotechnology and pharmaceutical R&D, coupled with supportive regulatory frameworks, is propelling the development and commercialization of miRNA-based products, positioning the market for robust expansion in the coming years.

Get Sample Copy Of This Report @ https://www.snsinsider.com/sample-request/3143

Market Scope & Overview

The scope, global demand, marketability, profitability, and potential of the industry are all extensively covered in the most recent market research report. The research report examines the market in depth and offers details on a variety of subjects, including market drivers, constraints, opportunities, and threats. The potential for growth at the international, regional, and industrial levels is examined by MicroRNA Market research. The study also examines the consequences of the epidemic and offers suggestions for reducing market volatility.

The market research report presents an international viewpoint on the level of industry competition in various regions and markets throughout the world. A dashboard analysis of well-known companies is also included in the MicroRNA Market research report, showing their successful marketing strategies, market presence, and most recent successes in both historical and contemporary contexts.

Market Segmentation Analysis

By Products & Services

Services

Type

Isolation & Purification

miRNA cDNA Synthesis

Profiling, Localization, & Quantification

Functional Analysis

Others

Specimen

Whole Blood

Serum

Plasma

FFPE

Fresh Frozen Tissue

Others

Products

Instruments

Technology

Real Time PCR

Microarray

NGS

Others

Workflow

Isolation & Purification

miRNA cDNA Synthesis

Profiling, Localization, & Quantification

Functional Analysis & Others

Others

Consumables

Specimens

Whole Blood

Serum

Plasma

FFPE

Fresh Frozen Tissue

Others

Workflow

Isolation & Purification

miRNA cDNA Synthesis

Profiling, Localization, & Quantification

Functional Analysis

Others

By Application

Cancer

Infectious Diseases

Immunological Disorder

Cardiovascular Disease

Neurological Disease

Others

COVID-19 Impact Analysis

A comprehensive risk analysis and business ideas for the target market were created over time. The market circumstances prior to and following COVID-19 are also contrasted in this study paper. The influence of the COVID-19 epidemic on the market was thoroughly examined by the MicroRNA Market report.

Regional Outlook

In order to obtain qualitative and quantitative market data from internal and external sources, extensive multi-level research was first done. The plan also calls for the development of projections and overviews of regional markets for each category. During the MicroRNA Market research, the total market size was calculated using both primary and secondary data.

Competitive Analysis

Genuine information can help investors make smarter financial choices. In order to assist companies in the MicroRNA Market sector in making wiser decisions, the research's competition landscape section also includes the most recent information on recent partnerships, mergers, and acquisitions as well as key competitors' tactics.

Key Questions Answered in the MicroRNA Market Report

What impact will COVID-19 have on your target market?

Which market considerations have recently influenced decisions the most?

What are the target market's opportunities, threats, and prospects for the future?

Conclusion

In order to provide a thorough picture of the industry and help businesses better appreciate the possibilities offered by the various regional regions, the research report examines the MicroRNA Market market.

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Begin forwarded message: > From: evoldir@evol.biology.mcmaster.ca > Subject: Job: California_InstituteTech.FunctionalGeneticTech > Date: 19 August 2023 at 06:54:20 BST > To: rdmpage@gmail.com > > > The Parker lab at Caltech is searching for a Functional Genetic Technician > to generate genetically modified rove beetles (Dalotia coriaria) for > the study of behavior and chemical ecology. > > The successful candidate will work with the Principal Investigator and > lab members (graduate students and postdocs) to establish and maintain > engineered beetle lines created using CRISPR/CAS9 gene editing and > transposon-mediated transgenesis technologies. > > The position requires competence in molecular biology techniques for the > design and construction of plasmid vectors and sgRNAs for insect genome > engineering. The position also demands careful screening and husbandry of > insect cultures. Necessary bioinformatic skills can be acquired through > training in the Parker lab. > > Essential Job Duties > > - Molecular biology methods (DNA/RNA extraction, PCR, cloning, cDNA >  synthesis). > - Design and synthesis of sgRNAs and transformation vectors for gene >  editing and transgenic manipulation of rove beetles. > - Screening and husbandry of transgenic and mutant rove beetles to >  establish stable genetically modified lines. > - Working with genomic and transcriptomic data for the above tasks. > - Maintenance of genetically modified beetles. > - Working with postdocs and students to design and engineer project- >  specific genetic constructs. > - Basic Qualifications > > - BS in a biology-related field with at least 1 year of relevant work >  experience. > - Demonstrated molecular biology experience. > - Preferred Qualifications > > Experience with insect genetic manipulation. > Experience with plasmid design and construction. > Experience with insect transgenesis. > Experience with insect rearing/husbandry. > Required Documents > > Resume > > Apply: > https://ift.tt/I7dJ8U1 > For more info contact: joep@caltech.edu > > Joe Parker, Ph.D. > California Institute of Technology > Division of Biology and Biological Engineering > 1200 E. California Blvd. > MC 216-76 > Pasadena, CA 91125 > > Tel: +1 626 395 8729 > https://ift.tt/iksoeX3 > > "Parker, Joseph"

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Molecular Biology Enzymes Kits and Reagents Market Size, A Forecasted Outlook for 2023-2030

The latest market report published by Credence Research, Inc. “Global Molecular Biology Enzymes Kits and Reagents Market: Growth, Future Prospects, and Competitive Analysis, 2022 – 2030. The global demand for molecular biology enzymes kits and reagents market was valued at USD 20.2 Billion in 2022 and is expected to reach USD 38.12854245 Billion in 2030, growing at a CAGR of 9.50% between 2023 and 2030.

Molecular Biology Enzymes Kits and Reagents Market encompasses an extensive range of products crucial for various molecular biology protocols, research, and clinical applications. This market segment includes enzymes such as DNA and RNA polymerases, reverse transcriptases, restriction endonucleases, ligases, and various modifying enzymes, among others. These enzymes play pivotal roles in procedures like PCR, qPCR, sequencing, cloning, cDNA synthesis, and restriction digestion. Accompanying these enzymes are specialized reagents and kits optimized for specific applications, ensuring accuracy, reproducibility, and high yield. The demand in this market has been amplified due to the burgeoning biotechnology and pharmaceutical sectors, ongoing research in genetics and molecular biology, and the rising prevalence of infectious diseases and genetic disorders.

Additionally, the recent surge in genomic studies and personalized medicine has further propelled the requirement for high-quality enzymes and associated reagents. Major players in this space are continuously investing in R&D to offer innovative, efficient, and cost-effective solutions. However, the market's growth can be constrained by factors like high costs of certain products and stringent regulatory landscapes. Overall, the Molecular Biology Enzymes Kits and Reagents Market represents a dynamic and vital segment in the life sciences industry, with its products underpinning many modern biological and clinical breakthroughs.

Molecular Biology Enzymes Kits and Reagents Market Key Growth Trends have been a subject of great interest and significance in the scientific community. These trends highlight the constant evolution and advancements in molecular biology research, particularly in relation to enzymes, kits, and reagents. Researchers are continually seeking innovative solutions to optimize experimental procedures and improve efficiency. The market offers a wide range of enzymatic products that cater to diverse applications such as DNA amplification, sequencing, cloning, gene expression analysis, and protein purification. These growth trends indicate an increasing demand for user-friendly enzyme kits that provide reliable results with minimal effort. Furthermore, there is a growing emphasis on environmentally friendly products that reduce waste generation while maintaining high performance standards.

Molecular Biology Enzymes Kits and Reagents Market Key Offerings-

1. Enzymes

DNA Polymerases: Essential for PCR, qPCR, and sequencing applications.

RNA Polymerases: Used for in vitro transcription processes.

Reverse Transcriptases: Central to cDNA synthesis and RT-PCR.

Restriction Endonucleases: Enable DNA fragment analysis and cloning.

Ligases: Vital for linking DNA fragments together.

Modifying Enzymes: Such as methyltransferases and phosphatases, crucial for modifying DNA molecules.

2. Kits

PCR and qPCR Kits: Contain all necessary reagents for amplifying DNA or RNA.

cDNA Synthesis Kits: Offer reagents for synthesizing complementary DNA from RNA templates.

DNA Sequencing Kits: Provide the necessary components for DNA sequencing procedures.

Cloning Kits: Simplify the process of inserting DNA into vectors for expression or analysis.

Genotyping Kits: Facilitate the identification of genetic variations.

3. Reagents

Nucleotides: Basic building blocks for DNA and RNA synthesis.

Primers and Probes: Short strands of nucleic acids essential for processes like PCR.

Buffers and Solutions: Maintain optimal conditions for enzymatic reactions.

Dyes and Stains: Enable visualization of nucleic acids.

4. Sample Preparation Products

DNA and RNA Extraction and Purification Kits: Help in isolating nucleic acids from various samples.

Gel Extraction Kits: Allow for the purification of DNA fragments from agarose gels.

5. Assay Kits

Gene Expression Assays: Facilitate the study of gene activity.

Mutation Detection Assays: Enable the identification of genetic mutations.

Methylated DNA Detection Kits: Used to study DNA methylation patterns.

6. Library Preparation Kits

Essential for next-generation sequencing, these kits provide tools to create libraries from DNA or RNA samples for sequencing applications.

7. Accessory Products

Positive Controls: Provide known references for various assays.

Stabilizers and Preservatives: Extend the shelf-life of enzymes and other sensitive products.

Browse 250 pages report Molecular Biology Enzymes Kits and Reagents Market By Product (Kits & Reagents, Enzymes, Ligases, Restriction Endonucleases, Reverse Transcriptases, Other Enzymes) By Applications (PCR, Sequencing, Cloning, Epigenetics, Restriction Digestion, Synthetic Biology, Other) - Growth, Future Prospects & Competitive Analysis, 2016 – 2030)- https://www.credenceresearch.com/report/molecular-biology-enzymes-kits-and-reagents-market

Molecular Biology Enzymes Kits and Reagents Market Regional Analysis-

1. North America:

Status: Dominates the global market in terms of revenue.

Drivers: Robust biotechnological and pharmaceutical sectors, extensive R&D activities, advanced healthcare infrastructure, and the presence of key market players.

Key Countries: The United States, owing to its heavy investments in biopharmaceutical research and genomics, is a significant contributor.

2. Europe:

Status: Second-largest market after North America.

Drivers: Strong academic and research base, especially in countries like Germany and the UK, paired with substantial public and private funding.

Key Countries: Germany, UK, France, and Switzerland, with their rich history in pharmaceuticals and biotechnology research.

3. Asia-Pacific (APAC):

Status: Fastest-growing region in the market.

Drivers: Rapidly developing biotechnology sectors in countries like China and India, coupled with increasing government funding in healthcare and research. The region's large population also drives clinical research and diagnostics, necessitating molecular biology products.

Key Countries: China, India, Japan, and South Korea.

4. Latin America:

Status: Moderate growth compared to other regions.

Drivers: Gradual growth in biotechnology and healthcare sectors, increasing public awareness, and rising government investments in research.

Key Countries: Brazil and Mexico are the leading contributors due to their sizable pharmaceutical and biotech industries.

5. Middle East & Africa (MEA):

Status: Smaller market share but shows potential for growth.

Drivers: Increasing investments in healthcare and research, especially in affluent nations like the UAE and Saudi Arabia.

Key Countries: South Africa, UAE, and Saudi Arabia. South Africa, in particular, has shown significant strides in healthcare research.

Molecular Biology Enzymes Kits and Reagents Market Future Outlook-

The future of the Molecular Biology Enzymes Kits and Reagents Market is anticipated to be vibrant and dynamic, underpinned by a series of technological breakthroughs and evolving healthcare paradigms. As the world increasingly leans towards personalized medicine, the demand for molecular tools that facilitate precise genetic and genomic analyses is expected to surge. Additionally, the rising prevalence of chronic diseases and global health crises, such as pandemics, further emphasize the importance of molecular diagnostics and research, consequently bolstering the market. Technological advancements, especially in areas like next-generation sequencing, CRISPR-Cas genome editing, and single-cell analysis, will likely shape the market's landscape, opening new avenues for diagnostics and therapeutic interventions.

Moreover, emerging economies are set to play a pivotal role, with their escalating healthcare investments and burgeoning biotech sectors. However, the market isn't without challenges. Regulatory complexities, high costs of advanced solutions, and ethical considerations in areas like gene editing will require navigation. Collaborations, innovations, and strategic mergers will be key for market players to thrive. In essence, the Molecular Biology Enzymes Kits and Reagents Market is on the precipice of significant growth, poised to make monumental contributions to science and medicine.

Why to Buy This Report-

The report provides a qualitative as well as quantitative analysis of the global Molecular Biology Enzymes Kits and Reagents Market  by segments, current trends, drivers, restraints, opportunities, challenges, and market dynamics with the historical period from 2016-2020, the base year- 2021, and the projection period 2022-2028.

The report includes information on the competitive landscape, such as how the market's top competitors operate at the global, regional, and country levels.

Major nations in each region with their import/export statistics

The global Molecular Biology Enzymes Kits and Reagents Market  report also includes the analysis of the market at a global, regional, and country-level along with key market trends, major players analysis, market growth strategies, and key application areas.

Browse Full Report: https://www.credenceresearch.com/report/molecular-biology-enzymes-kits-and-reagents-market

Visit: https://www.credenceresearch.com/

Related Report: https://www.credenceresearch.com/report/cannabinoid-biosynthesis-market

Related Report: https://www.credenceresearch.com/report/oncology-molecular-diagnostics-market

Browse Our Blog: https://www.linkedin.com/pulse/molecular-biology-enzymes-kits-reagents-market-share-demand-singh

About Us -

Credence Research is a viable intelligence and market research platform that provides quantitative B2B research to more than 10,000 clients worldwide and is built on the Give principle. The company is a market research and consulting firm serving governments, non-legislative associations, non-profit organizations, and various organizations worldwide. We help our clients improve their execution in a lasting way and understand their most imperative objectives. For nearly a century, we’ve built a company well-prepared for this task.

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Global Gene Synthesis Market value

Global Gene Synthesis Market value

Scope of Global Gene Synthesis Market:

The latest business intelligence report on the Gene Synthesis Market offers a comprehensive overview of the pivotal aspects pertaining to this industry vertical. It incorporates an accurate assessment of historical records, projections, growth drivers, opportunities, challenges, and restraints, among others.

This research literature fragments the industry in terms of (segments). It individually assesses each segment based on its scope and provides valuable insights on its top revenue prospects. By using proven research methodologies, this document further includes granular insights into the geographical landscape while providing a 360-degree outlook of the growth trajectory of the highlighted regions.

The study on Gene Synthesis Market further characterizes the competitive terrain by incorporating crucial data about the top industry players. It comprises of the detailed portfolio of each of the mentioned companies while deeply scrutinizing the major strategies adopted by them to enhance their global footprint. The crucial components such as product pricing, partnerships, mergers & acquisitions, collaborations, and major developments associated with each player are also unveiled in this report.

This report is specially curated to empower the existing players, stakeholders, and new entrants about the ongoing trends of this market which will allow them to make informed business-centric decisions.

Browse In-depth Market Research Report (300 Pages) on Gene Synthesis Market:

Gene Synthesis Market Companies:

Thermo Genewiz

Eurofins Scientific

Quintara Biosciences

ATD Bio Ltd.

Fisher Scientific, Inc.

OriGene Technologies, Inc

Bioneer Corporation

Atum

Integrated DNA Technologies, Inc.

BioCat GmbH

GenScript

Eurogentec

Twist Bioscience.

LGC Biosearch Technologies

Eton Bioscience, Inc.

Bio Basic Inc.

SBS Genetech Co., Ltd.

Merck KGaA

Others.

Regional Insights:

The regions covered in this Global Gene Synthesis Market report are North America, Europe, Asia-Pacific, and Rest of the World. Based on country level, the market of Managed security service is subdivided into the U.S., Mexico, Canada, U.K., France, Germany, Italy, China, Japan, India, Southeast Asia, Middle East Asia (UAE, Saudi Arabia, Egypt) GCC, Africa, etc.

Global Gene Synthesis Market Segmentation:

By Method

Oligonucleotides

Phosphoramidite Reaction Cycle

High-Throughput Array-Based Gene Synthesis Technology

Ion Semiconductor Sequencing

Nanopore Sequencing

Gene Assembly

Polymerase-Based

Dual-Asymmetric (DA) PCR

Overlap Extension (OE)

Polymerase Cycling Assembly

Thermodynamically-Balanced Inside-Out (TBIO)

Microchip-Based Multiplex Gene Synthesis

Others

Ligase-Based

Shotgun Ligation

Two-Step Ligation and PCR

Ligase Chain Reaction

Brick-Based

Recombinant-Based

Sequence and Ligation Independent Cloning (SLIC)

Transformation-Associated Recombination

BioBrick Assembly

By Type

Gene Library Synthesis

Custom Gene Synthesis

cDNA

Customized Coding Sequences

Genomic DNA

RNAi Constructs

Others

By Component

Product

Services

By Application

Research and Development

Diagnosis

Therapeutics

Others

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