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Plant Secondary Metabolism - Science topic

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I experimented the modified vanilin method and for the calibration curve i used cathechin but there was no result. All my samples were without color. Please help me.
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Which solvents can be used to extract bioactive compounds from Prunus avium (cherry fruit)? Its for food aplication.
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It seems there's a number of brands and companies to choose from, and no one I know has performed this test.  Abcam's kit seems to be the most widely used from the literature.  Do you have any brands to recommend or any secret tips to share?
Thanks!
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Hell, I am working with this subject and am to use Abcam's kit . I want to know what you used finally, and can you offer me your protocol .
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I am a MSc student of plant biology and I am looking for a researcher who either have an interest or that studies the differences in the secondary metabolism of plants under space conditions or stress in general that can be applied to space research (radiation, microgravity, hipoxia, drought, mineral nutrition, etc). I am also interested in researchers that work with molecular biology of plants under the same conditions.
I am asking this because I am trying to find a lab for my PhD. I have a great interest in the subject!
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I am currently working on a paper on secondary metabolites of brown algae  I am having a hard time finding a good reference for the Rf factors of the different compounds which are common in plants and algae (e.g. alkaloids, terpene). I only used thin layer chromatography (TLC) for the separation of the different secondary metabolites. Are there any references that you can suggest that I use for this paper? 
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Thank you all for your answers, however i have always thought that there were established standards (standard Rf  for each class of phytochemical) in relation to the stationary and mobile phase used. I am also disturbed because it is my thinking that one plant extract might contain more than one form of a given phytochemical. Taking this into consideration, it might be a little difficult to get pure samples of the phytochemical
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 I want to purchase metatitanic acid, where can I get it?  or prepare it in lab.
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Me as well,,,love me some over the counter Metatitanic acid (H2TiO3) ! Mess/me if you find some.
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phytochemistry, pharmacognosy, natural product, isolation of natural compound
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With the glycosidic group, those will be fairly polar. for silica, you may need to run dichloromethane/methanol (and you can go to 100% methanol if needed if using newer type B silica. One can even run methanol/water (HILIC) on silica.
C18 is also another choice.
Compound #5 in the app note link below was purified with C18
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I'm searching for a data base of plants with their respective secundary metabolites. Any idea someone? Thanks
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Hi, you can also try the follwing sites:
and these books:
Belitz, H.D.; Grosch, W.; Schieberle, P. (2009) Food Chemistry. Springer ed. 
Harborne, J.B.; Baxter, H.; Moss, G.P. (1999) Phytochemical Dictionary: A Handbook of Bioactive Compounds from Plants. Taylor & Francis Ed.
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I worked on two related plants. I want to identify that both plants contain the same secondary metabolites by taking a single metabolites.
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Dear Jitendriya, you may have a look into the folioing (attached link) article related to your query.
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Hi everyone,
Suppose one wants to elucidate the pathway of a plant secondary metabolite, or of a class of plant secondary metabolites (e.g. alkaloids) in a certain organ (e.g. fruit). How should this question be addressed? Where should one start and proceed? Which techniques can be used in each step, and for what reason? There is no need for details on procedures, just the general idea.
It may be a too general question, with perhaps some obvious answers, but it would be really useful for me, who have never worked in this field.
Thank you in advance.
Best Regards,
Gustavo
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Very informative attached links Mr. Abdulla
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I need some opinions in terms of gene expression and secondary metabolites for plants grown under artificial light. I need to know if there are any big differences in terms of (anti-oxidant, flavanoids, pigments and other secondary metabolites) between plants grown under aritifical lights and plants grown under natural sunlight. Opinions with relevant citations would be much appreciate.
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 Photosynthetic processes are often modified in plants grown under artificial lighting, because lamps do not usually mimic the spectrum and energy of sunlight. Agronomically, new lighting technologies such as LEDs have the potential to cover fluence and wavelength requirements of plants, while allowing specific wavelengths to be enriched, thus supplying the light quantity and quality essential for different phases of growth. The biomass and metabolic products of cultivated plants can therefore be modified. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3949401/
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I'm new in the area of plant secondary metabolism. So i wanted to know if there is any way to genetically express some specific secondary metabolites (Alkaloids, Terpenes, etc) on diferent microorganisms (Bacteria, yeasts, microalgae).
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I need a database or books for known compounds NMR data. I isolated some compounds and their NMR data something different for this case, I am thinking these compounds are the same type.
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You may find some spectra of relevance in these metabolomic databases
good luck
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I tried to separate column fractions by LH-20 but unable. I used a different percentage of chloroform and methanol or higher percentage of methanol to chloroform. All the case, I am unable to purify the compounds.
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try using a single solvent since mixture of solvents can cause your sephadex to swell differently, besides, dont forget to perform TLC for all your fractions and visualise under different conditions (normal light, U.V., acid stain etc)... run your fraction slowly and repeatedly until you get a single spot ... all the best...
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(Various literatures showing from 720 to 780nm)
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The best way to know the precise wavelength is to plot absorption spectrum of the product (coloured complex) and realize by your-self. Well, as I stated in my earlier response, the absorption maxima shifts based on the concentration of end product. If the concentration of product is below one OD, then the precise wavelength is 765 nm. If you modify any protocol, first standard curve with Gallic acid according to your modified protocol (or even any other published protocol) and plot absorption spectrum of the end product in reaction mixture to confirm the precise wavelength by yourself. 
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Hydrophobic compounds are solubilised in water by using hydrotropes. After extraction, recovery of the target compound is done by dilution. Does anyone have example of using hydrotropic extraction at industrial scale? 
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It can be feasible. However the dilution process to recover the phytochemical can be energy consuming due to the evaporation process needed for the solvent reusability. 
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which plants have considerable amount of this material? and what is it's role?
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Answer from Fuenzalida is interesting. 
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Plant material is normally extracted after shade drying and grinding. Pulverized material is then extracted through different techniques like cold maceration or through soxhlet apparatus. If someone wants to get crude extract of a pulpy plant tissue (e.g., Aloe vera leaf or Broussonetia papyrifera fruit), what will be processing steps as shade drying for such tissues is rather impossible?
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Huma, the deterioration of the fleshy tissue is possible, but it all depends on the particular analyte you are trying to isolate from the material. In Hawai'i aloe is used for a multitude of purposes from dermatological ailments to gastrointestinal conditions. The many processes of extraction is what helps to bring to light the proper phytocompounds. Shade drying, or curing of plant material can cause decomposition, but usually if proper care and observation is taken you are able to determine the half-life of your drying process. One thing that was a great help for me starting out was sampling analyses for various phases of drying and solvent extraction. A wet process may also be undertook as mentioned previously where you skip drying/dehydration and continue with your extraction process and complete with a vacuum filtration process and/or drying, but I find that, in the case of orchids, sometimes it is better to skip drying and prepare slurry solutions then vacuum filter via fine sieves in a duplicate or triplicate manner to rid of a limited amount of solvent or diluent and any moisture or additional fleshy tissue. If you are concerned about the loss of analyte in question, I would recommend not drying the material before or after and using water or other complementary diluent as a solvent or no solvent prior to filtration. Don't be afraid to get your hands dirty and make mistakes, as errors are the predecessor of scientific discovery. Now, another technique you may undertake for drying would be to use an agitating steam bath to evaporate some of the fleshy material under low heat, but this, like all drying techniques, may lead to the destruction of vacuoles and their associated compounds stored for use; alcohols, acetonitrile, ethanol, etc. are also great for evaporating water based moisture in tissue, but this will also disassociate chemical bonds. I would like to also apologize that I am unable to link any related research for extraction techniques as I am out of the country until mid-August.
Jason
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Ask about can be extract this component individually or combined from food or plant origin .
If i can i need the extraction method and determination
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1. Overexpression of petunia chalcone isomerase in tomato results in fruit containing increased levels of flavonols
SR Muir, GJ Collins, S Robinson, S Hughes… - Nature …, 2001 - nature.com
... FM6203, Unilever commercial variety) leaf disks according to standard protocols 16 . ... RNA was
isolated according to the protocol of van Tunen et al 16 . ... | ISI | ChemPort |; Hertog, MGL, Hollman,
PCH & Venema, DP Optimization of a quantitative HPLC determination of potentially ...
2. BMC Veterinary Research | Full text | Protective effects of (1 ...
by SM Dhiyaaldeen - ‎2014 - ‎Related articles
Dec 31, 2014 - The gastro-protective effects of HPTP chalcone was estimated ..... Gastric wall mucus was determined based on the modified procedure of Corne et al. .... Woods RJ: Proceedings: A method for the quantitative estimation of ...
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All chemicals obtained from sigma in powdered form
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Extinction coefficients are dependent on the measurement wavelength, and the solvent used.  Hence, there is no universal extinction coefficient.  I'm guessing your aim is to quantify these compounds, and it looks like you already have the standards, so there is no need to determine extinction coefficients.  You can quantify them based on standard curves.
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My present research is focusing on the evolution of plant secondary metabolites within a particular group of plants of medicinal importance. I have gone through a review of Wink which I am enclosing. As we all know, this phenomenon occurs because of convergent evolution. But to establish it in a scientific manner how should I proceed. Any constructive opinion is most welcome.
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Get in touch with Vicki Funk at the Smithsonian - she was the first phylogeneticist to approach this problem - begin your literature review with 
Seaman, F. C. and V. A. Funk. 1983. Cladistic analysis of complex natural products: developing transformation series from sesquiterpene lactone data. Taxon 32: 1-27.
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I’m planning to search new secondary metabolites from rice which will be phytotoxic. I read about various methods for identification and quantification of rice allelochemicals and would like some advice about recommended techniques. I'm really looking to find out what you'd consider to be the most efficient method as cost and time are limiting factors.
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Dear Dr. B.R. Rajeswara Rao and  Dr. Jun Murata many many thanks for your valuable suggestions. I will follow these link. However, Dear Dr.  Jun Murata, I am trying to identify and quantify allelochemicals from rice root exudates and from shoots therefore I am searching a method that I can learn how to make sample preparation for Chromatography (GC MS and LC MS) and NMR analysis.  If any please help me regarding this.
Finally, thank you for your nice cooperation.
Best Regards
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I made some ethanolic extracts, but I don't know which are the factors (temperature, pH) involved in vanillin degradation that may alter those samples before HPLC analysis. 
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Dear Ms Luna
Sorry
I do not have any experience of this sort of work
Dan Eisikowitch
Professor emeritus
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I need a place in Egypt has a headspace apparatus to analyze traces of volatile oil or aroma of plants
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Dear Marwa Emad,
For volatile oil without using an extraction solvent, Dynamic Headspace is preferable (http://www.cdsanalytical.com/instruments/headspace/why_headspace.html). As analytes are very volatile, they can be sampled, injected and then separated as gases (https://www.youtube.com/watch?v=Vwol2oxdJ7s). 
Headspace sampling is essentially a separation technique in which volatile material may be extracted from a heavier sample matrix and injected into a gas chromatograph for analysis. To appreciate the principle, let’s consider an application that is well suited for headspace sampling: perfume. The composition of perfume may be highly complex containing water, alcohol, essential oils etc. If we inject such a sample directly into a typical GC injector and column, a lot of time may be wasted in producing the chromatogram by eluting compounds that we have no interest in. Moreover, many of these compounds may not be suited to gas chromatography and will gradually contaminate the system or even react with the stationary phase in the column so their presence is not desirable.
Kindly enclosed the principles of the techniques and the difference between a static and dynamic headspace.  
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so anybody have any idia how to convert this non solid material to solid constituents ,i mean if any gentleman done this work for any plants can explain it how to separate this extracted to solid parts ,thanks for your answer 
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You will always get semi-solid extract. Physical characteristics of an extract depends on group of compounds present in the extract i.e. If saponins are present than you will always get gummy type semi-solid. Simple way to get solid particle is after removing solvent, dry the extract on anhydrous calcium chloride in a vaccum desiccator.
However, some worker processes the extract with starch or sugars prior to evaporation of solvent to get free flowing particles.
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Which is the best method to separate and characterize compounds from Crude plant extracts (Polar and non polar solvents )?
Please suggest different techniques.
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there is no general method for isolation of all classes of compounds. however it depends on your approach of isolation either you are targeting certain class of compounds or you are doing bioassay guided fractionation or even aiming to study metabolomics.
in the first case the extraction method depends on the chemistry of the target compound,for example; if you aim to isolate alkaloids you can perform acid base extraction, if you aim to isolate hydrocarbons, non oxygenated terpenes or sterols you can carry out saponification of the least polar extract (hexane). for oxygenated terpenes the chloroform extract is suitable. the ethyl acetate extract contains a large variety of compounds, the butanol extract contains polar compounds like saponins.
meanwhile miscellaneous examples can happen
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Hi, I am doing derivatization of luteolin (standard) for GCMS analysis. My derivatization reagent is BSTFA and TMCS (99:1 ratio). I tried diferent temperature (60, 70,80,110) and time (15min, 30min,45min,60min), but still cannot find my luteotin. Can anyone give me any suggetstions?
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I tried as well this way (derivatization with BSTFA wih GC-MS analysis) of kaempferol, myricetin, syringetine. I think it's not a problem of derivatization but analysis choice. I never succeed to see these derivatized flavonoids.
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I am doing derivatization of  different flavonoids (Quercetin, morin, rutin, luteolin, apigenin-7- glucoside, luteolin-7-glucosie, catechin, taxifolin) using BSTFA and TMCS reagent (99:1 ratio) for GC/MS analysis. However I got always a peak which does not correspond to my flavonoid analyte, where the NIST library identifies it as fatty acid.
I do the derivatization as follows: the standard is dissolved in minimum amount of methanol, then ethyl acetate is added, followed by the derivatizing agent . the resulting solution is then heated at 100 C for 90 minutes, and then one micro liter is injected into the GC/MS.
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Hi,  i also come into such problems and I also use BSTFA and TMCS reagent (99:1 ratio). Have you found your flavonoids in GCMS? What is your conditions and derivatization reagent?
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Glucosinolates; plant secondary metabolites are known to be very effective against cancer, but some reports suggest that they are also toxic if used as primary food source. Does any researcher has any published material on toxicity limits of these compounds?
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Hi, Saeed,
Ref.:  Das, S., Srinibas, Tyagi, A. K., and Kaur, H. (2000). Cancer modulation by glucosinolates: A review. Curr. Sci. 79: 1665–1671. 
According to Das S et al and Lavecchia T et al, overdose of glucosinolates can cause thyroid dysfunction causing hypertrophy and goiter. 
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Such as lectin and saponin
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Respected Sir,
As Olukayode Solomon Ajayi Sir said, you have to perform qualitative preliminary phytochemical screening followed by HPTLC, GC-MS/LC-MS analysis and finally extraction, isolation, purification, identification and structural elucidation of the phytochemical constituents (compound isolation).
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The non-treated material is comparatively soft and cannot be cut to make a thin section (about 30 μm thick). And I want to prevent several seconfary metabolites (water and alchol soluble) dissolving from it by various embedding treatments.
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Depending on the tissue, you should be able to make a few sections of an appropriate thickness by hand. It takes practice, but I would start by trimming down a chunk and placing it on a slide, with water to keep it from drying out. Then you can place another slide across it as a guide (like a carpenter's fence). Using a fresh razor-blade make your first cut with the blade edge vertical; and then, not moving the fence, make a second cut with the blade edge angled in slightly. With practice you can produce a nice section in which the thickness varies across the width of the section from very thin at what was the top of the specimen to somewhat thicker at the bottom - wedge-shaped. Once you get the technique down it is quite easy to control the thickness nicely. It's true that the thickness is not uniform across the section, but for most microscopical observation at 20X or higher, the field of view is small enough so that you don't really notice the thickness difference. Works well for herbaceous stems, some leaves, and even stems with some secondary wood formation, as long as the blade is sharp. Use a fresh blade every time! You need to trim the specimen carefully so that it lies flat on the cutting surface (the first slide) and make sure your axial orientation is precise so that you get true cross-sections.
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Anticancer activity of medicinal plants.
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There are many reports that ROS activate NF-kB activation and subsquently induce ATF3 transcriptional activation, which is associated with the apoptosis of human colorectal cancer cells.
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I would like to quantify some of phenolic and flavonoid compounds by HPLC. How can I extract these compound from plant and what is the solvent of HPLC with what program and percent of solvent?
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(1) I am using 50mg dried sample + 1ml Methanol, sonication for 60 minutes at 50'C, filter with 0.45um membrane filter- 10 ul injcet to HPLC
(2) For the solvent : A=0.1 % Formic acid B=Acetonitrile
Time Flow (ml/min) %A %B
0 1.0 85 15
12 1.0 75 25
22 1.0 75 25
25 1.0 85 15
35 1.0 85 15
PDA detector
Gud luck
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I am looking to extract hesperidin, a known flavanone glycoside, from J. sambac roots. However, I would like to know if there are any other techniques I can implore in order to further increase the yield and also increase the yield's purity. Any help would be greatly appreciated.
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There are a variety of methods and strategies employed for the extraction of a particular class of flavonoid. Flavonoids (particularly glycosides) can be degraded by enzyme action when collected plant material is fresh or non-dried. It is thus advisable to use dry, lyophillized or frozen samples. The plant material is ground into a powder when dry material is used. Another important consideration is polarity. Isoflavones, flavanones, methylated flavones and flavonols are less polar flavonoids and are extracted with organic compounds like chloroform, dichloromethane, diethyl ether or ethyl acetate. Flavonoid glycosides and more polar aglycones are extracted with alcohols or alcohol water mixtures. The bulk of extractions of flavonoid containing material are still performed by simple direct solvent extraction.
Powdered plant material can also be extracted in a Soxhlet apparatus, first with hexane to remove lipids, and then with ethyl acetate or ethanol to obtain phenolics. This approach is not suitable for heat-sensitive compounds. However a more convenient and frequently used procedure is sequential solvent extraction in which the powdered raw material is first extracted with dichloromethane, for isolation of flavonoid aglycones and less polar material. A subsequent step with an alcohol will extract flavonoid glycosides and polar constituents. Flavanone solubility depends on the pH of water-containing solutions. Flavan-3-ols (catechins, proanthocyanidins and condensed tannins) can often be extracted directly with water. However, the composition of extract does vary with the solvent- whether water, methanol, ethanol, acetone or ethyl acetate.
Hesperidine has been extracted from a variety of sources using both analytical as well as preparative techniques. These techniques are very easily accessible on the internet in the form of freely available research papers, hence I am not giving the details of that procedure as you can access them yourself. I am attaching one of them for your perusal.
Regards
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Does anyone use "pESC Vectors (pESC-HIS, pESC-LEU, pESC-TRP and pESC-URA )" for protein co-expression in S. cerevisiae?
I need them for my co-expression studies.
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I have worked a lot with this vectors and my first advice is to carefully read the information brochure to choose the best one. Besides, depending on the size of your target gene it will be easier or harder to clone it into these vectors.
Once cloned and transformed in E. coli I recommend to you to use a high concentration of Amp when growing your culture for DNA amplification (maxi or megaprep) since the yield is not very high.
For the transformation in yeast I recommend to you the lithium acetate method which is easy, cheap an fast.
Another hint is that when you grow your cultures, this time yeast, in order to obtain your proteins, you should start with a small inoculum in your deficient media with a 2% glucose (v/v) for an overnight. After that you can do a 1/1000 dilution in the presence of 2% Rafinose. This is very important as you have to eliminate any trace of Glc because it is a repressor of GAL promoters. After this o/n growing in the presence of rafinose you can "start" your protein expression with a "great population in a very good mood" by adding 2% galatose final (v/v), ca. 6h is the optimum for me.
I hope you find this highlights useful. Best of lucks!
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Does anyone have experience with detergents for a solubilization of membrane proteins that conserves a specific membrane-protein sterol interaction?
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good point! Thanks for the answers Maciej ! Cheers ML
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Is there specific elicitors to increase specific compound or a general elicitor is enough to increase all the secondary metabolites? How can one choose elicitors? Is it plant specific, desired compound specific or something else?
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Thank you Mr. Henrick and Gehan amin for your valuable suggestions....