Transgenics - Science topic

I recently started working with zebrafish and confocal microscopy. My samples are double transgenic for gfap_mcherry and pcna_gfp. gfap labels Müller glia in the retina, and pcna is a proliferation marker facilitating DNA polymerase during proliferation. I should start imaging at d3 using multiple confocal microscopes to decide the most suitable one but I got very diffused gfp signal with all microscopes. What can be the reason for that?

I would like to known what is the hygromycin concentration usually used in selection media to select and regenerate transgenic MicroTom tomato plantlets.

I've recently come across the method of agroinjection and vacuum infiltration of various plant tissues (seedlings, potato eyes, tomato fruit). These methods are new to me and very fascinating but I've been having a hard time finding answers regarding these methods.

1) Are these methods only for transient expression? Are the chances of creating a chimera, which then will produce transgenic seeds, extremely unlikely? Of course I would like to establish a stable transgenic line but transient expression might do for the experiment I have in mind.

2) Would multiple injections or vacuum infiltration sessions result in multiple integrations into the host genome? The outcome would probably still be a chimera, but I would think that multiple transfection rounds could greatly increase transgene expression, integration events and maximize the chances of producing seeds for a stable transgenic line. Is this a valid train of thought?

3) If my transgene encodes for a His-tagged protein and I am really only interested in purifying the His-tagged protein, transient expression alone can satisfy my needs. Is there an certain plant species or explant which is ideal for the production/purification of protein?

Thank you so much for any thoughts or suggestions while I continue to investigate this subject.

Hello,

Apologies if there's an obvious answer to this, I'm a first year PhD student and this task is entirely new to me. I'm including a lot of detail to convey my question clearly.

In D. melanogaster, the QF system is used to regulate the expression of transgenic constructs inserted within the genome of a specific fly line (similar to the UAS/Gal4 system).

In this system, the protein QF acts as a transcriptional activator, where in its active form it binds to a QUAS promoter and recruits transcriptional machinery.

One can easily obtain a fly stock which expresses QF from a stock centre like Bloomington. However, I need to design my own vector which will carry an insert that looks something like this:

Tub > QS - P2A - QF

Tub = Ubiquitous tubulin promoter, QS = Repressor of QF, P2A = "Self=cleaving" peptide, QF = Transcriptional activator

For this, I need to obtain the sequence of QS and QF; I have already obtained the former as its wild-type form is sufficient and easily accessible in NCBI.

However, I need a specific modified version of QF designed by Riabinina et al., 2015.

The modified QF protein, QF2w, is less toxic than the wild-type protein and is more robustly repressed by QS.

Is it possible to use online resources to obtain the sequence of QF2w and include it in my vector design? Or is my only option to contact the authors that designed this modified protein?

Best,

Panos

When we transfer plasmid in fungi for developing a transgenic fungi then how we can know that the fungi which grow on the media plates are transgenic or not?

Can we screen out fungi by adding antibiotics like Ampicine or kanamycine which is present in the plasmid?

A thin layer resembling a white callus was formed on the stem of a tomato plant transplanted from rooting food to soil. Is this dangerous for plant growth? No such situation was observed in the control plant.

Use of auxotrophic mutants lines for transformation of fungi is a traditional practise but it does not enable future discrimination between the wild type and transgenic line. Could basta gene serve to enable selection of the transgenic line from the wild type or from the back mutants of the transgenic line?

I need to check copy number in my transgenic rice plants. I am not able to get radiolabeled nucleotides here, so I have to buy a kit for southern blotting. Is these kits can also be used for doing EMSA?

Is it possible?

We have numerous previous articles and specific protocols by which you can regenerate transgenic rice plants. However, following the existing methods/protocol, if you do not get the positive outcome, I.e., no transgenic plants, what would be the possible reasons behind the failure to produce transgenic ones?

Hi, I did a quantitative analysis for my transgenic and control plants. The statistical analysis (p<0.05, One Way ANOVA, Turkey) showed that there are no significant differences between both plants. May I know how to indicate that data on the graph? Should I use the same alphabet on both plants to indicate it? Or there are other ways to indicate it?

Triple-gene Transgenic Cotton Exhibits Tolerance Against Insects and Herbicide.

Hellow fellow academics

I am currently in a dilemma and I would really appreciate some suggestions/guidance on the matter.

I have overexpressed my gene of interest (GOI) from wheat in Arabidopsis using the floral dip method and with strict screening on MS-Hygromycin, obtained my T3 transgenics. Now the problem is that while the selection, on the media has been successful, I am not able to get a band of my GOI on agarose gel after doing semi- RT-PCR. Initially, I thought that maybe my overexpression was unsuccessful so I took the T3 seeds to screen again on the media, but, the result was the same; the overexpression was successful and met the segregation ratio requirement of 100% germination. As this is my first time working with transgenics, please enlighten me on where I could be going wrong.

Please advise. Thank you for your time in advance.

Dee

Because expression has not always linear relationship with phenotype

We are considering different brands of stereo microscope to set up a small C. elegans laboratory. Looking for recommendations and any general advice on microscope features that you feel are important for C. elegans culture and research.

Features I expect are critical in deciding on the microscope we choose:

- Micrometer eye lenses - helpful for selecting/screening larval stages by size?

- Fluorescence capability - we won't need this at the start for wild-type strains, but I expect this is going to become a critical feature for transgenics. Have looked at the Nightsea fluorescence adapter, but leaning more towards an integrated system.

- Optical quality - we are looking into live worm tracking software - Is this possible with cheaper stereo microscopes or does the quality of the optics play a critical role in the software tracking accuracy?

- Ergonomics - C. elegans culture requires a lot of time on the scope - what precautions do you take for reducing neck and eye strain (e.g. is it best to have a scope with adjustable/tilting head?)

Thanks for your responses!

I have been thinking that I will just use EGFP to know whether my attempted transfection was successful, but I've been coming across some publications that say over expression of GFP can cause cell death. I need to create a stable line of transgenic organisms that have rather delicate embryos, and the organisms themselves are mildly fluorescent, so I am concerned that I need to hit a sweet spot of fluorescence that may be difficult to achieve. Too much and my cells die and too little and its not detectable. Does anybody know of another method that doesn't require expression of fluorescent proteins and doesn't require me to kill my organisms for sequencing? Thanks.

For western blot sample preparation of mice brain tissue (control and transgenic groups), I used RIPA with stainless steel beads for homogenization. After centrifugation, transgenic samples had a muddy pellet of tissue with steel beads, but control only had the beads and no visible tissue. I used the supernatant of both groups for BCA assay.

Protein concentrations from control group turned out to be great (at least 10 mg/ml). Transgenic tissue concentrations on the other hand were almost 10 times less. Around 1 mg/ml or under. Obviously, it's because the tissues did not get disrupted as well as the control tissues. I repeated the procedure for transgenic samples, nothing changed.

Any ideas and help would be much appreciated. Thank you!

Edit: I am attaching my sds-page gel photo to give an idea of how different the results come out. First four samples are control, last four are transgenic. Each sample is calculated to be 20ug.

Hello seniors! I am stuck in my work now I really need your help. I have a transgenic rice crop. For this, I got positive bands for my transgene in T1 & T2 generation plants. But in Generation 3 I am not getting any single positive band.

What I did do? I did the gradient PCR to check the correct annealing temperature. In my PCR I am getting positive bands for plasmid and for housekeeping gene but no bands for the transgene. Is it Gene's escape? or some other reason? Kindly guide me in this.

Your suggestions will be appreciated.

For transformation purpose

I constructed the transgenic vector, which had no problem in transiently expressing the protein first, and then transformed the Arabidopsis thaliana by dipping. The positive lines were screened by resistant medium first, and the DNA of T1 generation plants was extracted. The PCR identification results were correct, but the protein expression could not be detected by Western.

Hi! I need some help please guide me. I have Transgenic rice seeds. For PMI gene the PCR results are positive. But when i germinate that seeds on PMI selection media no germination occurs. What its means? either PCR gave false positive result or my seeds are not transgenic?

Hello everyone

I want to extract and isolate a plasmid from transgenic worms. Is there a specific procedure or a protocol for this purpose?

Thanks

Currently, I am working on detection of transgenic genes in corn plant, but I am facing many problems. For example, in some primers, the non-transgenic (control as witness) sample gives band as a PCR result while in some primers it doesn't. I have to add that we must not see a band in the non-transgenic sample.

What can be done to eliminate non-transgenic pollution?

The second question is that we do the negative control test with utmost care, why do we regularly see water contamination in some primers such as DEP98? What temperature and time cycles do you recommend to avoid such mentioned problems?

why do we use negative selection marker genes in transgenic plants as they won't survive?

and can we grow transgenic cells by negative selection?

I treated them with 70% alc, 2.5% NaOCl, 0.1 HgCl2, and 1g/l bevastine. These chem were used in diff combinations and for diff time periods. Infection still occurs. Please suggest any other way of disinfection and sterilization.

There are only few reports on the development of transgenic and tissue culture manipulations in indica rice. What could be the genetic, molecular, physiological reasons? which make indica cultivars to be more recalcitrant to tissue culture regeneration and transgenic developments.

Phenomena: In a recent study, RLB (RICE LATERAL BRANCH) gene were demonstrated to control the plant height of rice. A loss-function mutation of the gene led to reduced stature. However, when the authors overexpressed the gene, the transgenic lines also showed reduced plant height. The authors further determined the expression levels of endogenous and transformed RLB copies, and found that both of them were decreased to very low levels.

Question: RLB gene was proofed to positively regulate the plant height of rice, we expected that overexpression of RLB may cause overaccumulation of the RLB gene transcription and increase to some extent, at least not reduce the plant height.

Why overexpression of RLB resulted in reduced transcription for both endogenous and transformed RLB copies and caused the similar phenotype with the mutant?

Based on the knowledge we have learned about small RNA biogenesis and functions, please:

1) give the possible explanation of the phenomena;

2) describe the mechanism may cause the above phenomena;

3) design experiments to proof your hypothesis.

Hello,

I work with a knockout model infecting cells with cre-adenovirus to induce the deletion.

I am analyzing my qPCR results and I am not quiet certain about the best statisticalmethod to use, because I want to analyze the correlation of cre-adenoviral infection and animal genetic status (basically, I want to know how much of the gene expression is affected by cre-adenoviral infection by itself and how much is as a result of the mutation).

To evaluate that I have four groups:

- Wild-type cells not infected

- Wild-type cells cre-infected (n=3 for wild-type cells)

- Transgenic cells not infected

- Transgenic cells cre-infected (n=6 for transgenic cells)

I have tried different statistical models, but I believe I am still lacking understanding about choosing a statistical method.

PS: the reason for sample size differences was the unexpected result of gene expression being significantly affected in wild-type cells infected with cre-adenovirus - when I proved that wild-type cells were also affected, I did not have the opportunity to gather more samples.

Thank you

In a recent study, RLB (RICE LATERAL BRANCH) gene were demonstrated to control the plant height of rice. A loss-function mutation of the gene led to reduced stature. However, when the authors overexpressed the gene, the transgenic lines also showed reduced plant height. The authors further determined the expression levels of endogenous and transformed RLB copies, and found that both of them were decreased to very low levels.

Question: RLB gene was proofed to positively regulate the plant height of rice, we expected that overexpression of RLB may cause overaccumulation of the RLB gene transcription and increase to some extent, at least not reduce, the plant height. Why overexpression of RLB resulted in reduced transcription for both endogenous and transformed RLB copies and caused the similar phenotype with the mutant? Based on the knowledge we have learned about small RNA biogenesis and functions, please:

1) give the possible explanation of the phenomena;

2) describe the mechanism may cause the above phenomena;

Our lab uses tissue culture to maintain populations of Populus trichocarpa Nisgually, Populus tremula x alba INRA 717-1B4, and transgenic Populus deltoides x nigra plants (created via agrobactrium-mediated transformation). Recently we have noticed what appears to be callusing and sometimes leaf formation on the basal part of the stem in our fresh subcultures (see pictures), which then do not form roots. This problem has emerged within the last month, although some more established rooted cultures also have small leaves growing on their roots or out of the parts of the stems submerged in media. The media itself doesn't appear to be the cause, since the callus phenotype is only present in a subset of plants cultured in media prepared on a given day. We have also observed callusing in plants from all three populations, and it seems possible that the causative agent may be being passed on from older plants to fresh subcultures or between plants that are being subcultured from on the same day. There are also no visible signs of wounding or contamination in the affected plants. If anyone has seen or dealt with a similar issue, your input is much appreciated. Thank you!

If a transgenic mouse expresses transgenic T cell receptors that recognize an antigen derived from E. Coli presented by an MHC-class II allele expressed by the transgenic mouse, what is the likely scenario for this transgenic mouse?

How do you justify the usage of antibiotics in PTC at the time of

1) explant sterilization, or 2) within the plant tissue culture media.

is it suggestable/ethical to use antibiotics in normal tissue culture experiments? (not for the transgenic selection etc)

if yes how?

if not why?

I have applied initially the one-way ANOVA followed by TUKY and Kramer. May I ask you to verify if the method I used is appropriate for the data and advice?

I have 10 different transgenic arabidopsis lines (plants)  containing viral protein; hence I have 10 different Genetically modified plant lines. These 10 are compared to a standard plant that has not been GM, i.e., lacks the viral protein, companions between the different transgenic lines, and responses of each line at each of the concentrations.

Statistical differences in the seedlings' total length between the different tested Arabidopsis lines under different hormone concentrations were assessed by a one-way analysis of variance (ANOVA) using Statistical Analysis System – JMP (SAS JMP®, 14.0.0; SAS Institute Inc., Cary, NC). If the ANOVA implied significance (p ≤ 0.05), then the means were tested with Tukey-Kramer Honestly Significant Difference post hoc test to determine the differences.

Perhaps this is a naive question but I'm relatively new to flow and am trying to learn about the correct way to gate. I am using bone marrow chimera mouse model to assess peripheral immune cell infiltration into the brain. GFP+ bone marrow was used to reconstitute non-GFP recipient animals, so GFP+ cells in the brain are indicative of cells from the periphery. I am interested in having a very general idea of which groups have the most infiltration (i.e. %GFP of parent population) and have the raw data that I'd like to gate myself.

My question is, can the exact same gating be used for each sample? They are different genotypes and done across different days, but they are virtually indistinguishable from each other and in my mind it makes sense to have a consistent definition for what I'm considering a singlet or GFP+. I'm gating out debris, then single cells with FSC and SSC, then gating GFP+. No antibodies are used here, just the GFP from the transgenic line. Thanks!

Hello all, I am wanting to grow sorghum seed on skoog media with antibiotic to begin a genotyping process of transgenic lines. The seed is mature. If this is possible, would anyone be able to provide a protocol? If this is not possible, is there any other possible method that would work?

Thanks in advance!

Hello, everyone. I used a transgenic vector for SCNT and successfully obtained cloned individuals. I screened the cloned individuals for the presence of vector by PCR and distributed individuals into 2 groups: Group 1 containing transgenic vector and Group 2: without transgenic vector (called control). Next, I performed QPCR to evaluate the expression of gene. I found individuals with transgenic vectors (Group 1) also had overexpression of gene. But there were also few individuals in group 1 that although contains a transgenic vector but the expression of the specific gene was similar to the control group (without vector). So, need an expert opinion that why the few individuals who presented normal gene expression was with transgenic vector?

Thanking you in Advance.

Kind Regards

Which approach is better for in vivo calcium imaging in the brain with miniscope: image in virus-injected (AAV GCAMP6) or transgenic GCAMP6f animals?

Give three examples where this method is/was used. May you please send links of journal articles.

Dear colleagues,

for my research purpose i want to cross a transgenic GFP-reporter strain with a mutant strain. What is the optimal procedure to obtain a strain which inherits the transgenic genotype as well as the allel of the mutation? Can anyone provide a very detailed protocol to proceed the crossing including the observation of the transgene and the single-worm PCR?

Best regards

Fabian

I'm trying to buy a Vibratome VT-1000S. What are other brands and model similar to this one. Purpose is to section Medicago truncatula nodules.

In the modern precision breading, CRISPR cas 9 techniques of gene editing (deletion, insertion) is taking over the transgenic genetic engineering (GE) of crops and animals of importance to humans. At this point is there any reason for us to worry about GMO safety or adverse environmental effects? Why or why not? Please share us you opinions with supporting literatures.

The transgenic gene is suspected to be within another gene on mouse chromosome 1. Is there any indication that I can use in order to design a pair of primers? The objective is to ultimately find the location of this inserted gene in order to determine the allele combination of each mouse subject, being homozygous or heterozygous.

After Y1H screening, I got several TFs which can bind with the promoter of my gene. So I want to figure out if these TFs are activators or repressors to my gene. For the next step experiment, do I need to make the stably promoter::EGFP transgenic lines or conduct Dual Luciferase reporter assay? Although my advisor suggested me to make the stably promoter::EGFP transgenic lines, I am a little bit confused why I need the stably promoter::EGFP transgenic lines.

How to find the candidate genes to validate their role through functional genomics experiments such as cloning, transgenics, over-expression, localization, and its interaction with other proteins and DNA, etc.

1)Do we need to study a lot of literature and see which genes role is not deciphered in particular traits e.g. drought stress?

2.) Do we need to perform our own transcriptome or comparative genomic studies or analyze already published studies from literature?

3. ) Do we need to perform our own marker traits association(QTLs) study or already published studies?

4.) Some people functionally characterize already known genes(say arabidopsis) to plant of their interest (legumes). But is it a significant or novel research problem to work upon?

5. all of the above.

Many of my colleagues are trying to improve stress tolerance in stress-sensitive crops through a transgenic approach, because of climate change and the potential to use marginal lands. However, there has been very little progress in the applied aspect of it. We are also involved in a similar project and cloned 25 genes from various sources to test their potential to improve stress tolerance in yeast and Arabidopsis some of them were also transformed in eggplant and in tomato, generated several transgenic lines. Some of these genes have already been shown to confer stress tolerance in various publications, such as NHX1, TRK1, HKT1, etc., however, none of them produced any significant results in our lab. It turns out that stress-mediated reprogramming resulted in draining transgenic plants valuable resources by executing a stress response, even though the transgene imparted stress resistance trait. Our initial data suggests that the use of PGP microbes can improve environmental fitness by overriding the stress-mediated reprograming constraint, thereby suggesting a combined approach might be more practical and applied.

I would like to initiate an interactive discussion in this regard so that we can move this transgenic approach towards practical application.

I established the stable cell lines to express the protein driven by CMV promoter. It seems CMV promoter is easy to lose the expression of transgenic gene after several times cell passages. Does anyone suggest which promoter is the best to establish stable cell lines in colon cancer cell lines? 

I have overexpressed an algal gene in a grass crop. I did the qRTPCR for the gene and can see that the gene is overexpressing in transgenic lines by looking at the result. But since this is not an endogenous gene I cannot normalize the value for non-transgenic as 1 in the double delta Ct method. I have the Ct values for the target gene and the housekeeping gene. I tried the fold change method (Housekeeping Ct- target Ct) but that did not work for my case. I need to find a method to analyze my data.

What are the conditions for the vonversion? in many countries where conventional agriculture predominates, it seems something very difficult or impossible to achieve, I am referring to the conversion of large-scale conventional systems to agroecological systems.

I would like to know your opinion or experience, because I only know small-scale agroecological productions.

Genotyping PCR is used to determine the genotype of an organism (e.g., WT vs. mutant, or WT vs. transgenic)

Hello,

Which of these genetically altered organisms is more suitable for Scientific research? Is it the Transgenic hybrids or the Mutagenic organisms? Which is more genetically stable to be used as vectors for genome editing of plants?

Thank you in advance

I found the morphological mutation on my CRISPR/Cas9 transgenic potato tuber, in detail, which didn't enlarge ,was tiny and highly branched (secondary grew?), so that I want to know whether there are potato mutants of which phenotype is similar to my mut-tuber.

I am planning to sort transgenic hESC lines based on reporter lines fluorescent markers, to see if FACs sorting of progenitors results in greater Da yields. For my control study I have included a scrambled gRNA subculture. I wondered if I would have to make all subcultures, even those which will not be sorted (for comparison of sorted vs unsorted) reporter lines as a control?

Alternatively, if this is not necessary i will just compared transgenic sorted culture outcomes with original hESC?

I have two sets of qPCR data taken from the same sample type at two different points in time: one was acquired by researcher A two years ago with parameters A (Ta = 57 ºC, etc.); the other by researcher B recently with parameters B (T = 59 ºC, etc.). The data compare expression levels of a single gene between wild-type and transgenic animals. Reconciliation of these two sets would avoid further animal experimentation and increase sample size. Is there any way I can peacefully do this without feeling guilty? Thanks.

I'm trying to do genotyping of mouse DNA by the use of qPCR.

I'm using HTRA1 primer and GAPDH primer and SYBR green.

My problem is that I do not see a difference between my transgenic and the wildtype? I would expect a difference in the HTRA1 gene between the transgenic and the wildtype.

Anyone with the same problem?

I am interested to know the whole procedure for segregation including population size for segregation analysis to get homozygous transgenic plants.

Dear all,

I am trying to find an alternative to southern hybridization for copy number detection in transgenic Arabidopsis. I have attempted the QD PCR protocol (T.Kihara ) for the protocol but I am unable to really understand its rationale.

If anyone can provide me an alternative for copy number detection or provide me a simplistic reasoning behind the gel images from QD PCR, it would be of great help.

Thank you!

I have transgenic lines of arabidopsis expressing aequorin in the tonoplast, and am using them to determine calcium fluxes in week-old seedlings after certain treatments. The issue I am running into is that I am not sure if I am normalizing my readings from the luminometer to the summation of all the luminescence readings taken during the experiment, or only after discharging the aequorin with 2M CaCl2 in 20% EtOH. And if so, do I integrate those readings with respect to time or simply leave it as the sum? Thank you for any help, I appreciate it!

Hello everybody,

I would like to create a transgenic line for several proteins using a fusion-protein approach with Halo, SNAP or CLIP-tag. However, I cannot find a single paper where any of these tags have been fused with any protein to make a transgenic line.

I am mostly worried if how the ligand cannot reached the nucleus and thus the fusion-tag protein. I have read in mammalian cells, they are immersing cells in a bath containing the tags, that traverse the cell membrane and can find its target.

Does anyone has experience with these tags genetically encoded in model organisms?

Thanks a lot!

Noemie.

I want to make 10 transgenic Arabidopsis lines. After I collected T0 seeds, I stored them in room temperature for about 1 week. I sterilized the seeds with 70% ethonal alcohol for 7 min. The plates were dark and 4 degree for 3 days. Then they were transfered to white light. However, after 7 days, 2 cases of seeds were about 100% germinated. The other 8 cases were about 10% germinated. How could that happened? I don't think the seeds were died during I sterilization.

I am using rice variety Taipei 309 for generating transgenic. I have collected T0 seeds and want to know how long are they viable?

Thanks

I have been trying to generate transgenic poplar plants NM6 in WPM media and with different rooting hormones. My plants haven't been rooting so far.I transferred some into 1/2 MS with IBA, but there are still no roots. I welcome any suggestions you might have!

I am trying to isolate total protein from transgenic brassica roots but the samples are getting degraded due to proteasas so could anyone tell me what is best way to inhibit these proteases and isolate total protein from roots?

I got first generation of transgenic line, and did PCR screening. but when i inoculated PCR positive seeds to antibiotic plates for segregation analysis, I am not getting the mendelian Ratio. Can anyone please suggest me, what to do and how could I screen the transgenic plants?

I can get good hairy roots by Agrobacterium rhizogenous, but when I infect explants by A.rhizogenous that contain recombinant plasmid, I couldn’t get any hairy root. How much is possible to not get hairy roots and express a transgene coincidently in the infection by A.rhizogenous. In other words, if I transfer recombinant plasmid into A.rhizogenous there isn't any hairy root. But if I use A.rhizogenous by itself without any change, I see lots of good hairy roots.

Why is it difficult to grow Arabidopsis thaliana from callus (Single cell/transgenic cell)? Are there any technical limitations (if yes, then what)?

On the other hand, it is easy to grow other plants from callus (e.g. Brassica Napus).

Hi all, I have to give some low Nitrogen treatment in ms medium plates to my transgenic tobacco seedling, but i don't know how to prepare Ms modified plates with low Nitrogen.

As 1/2 ms medium powder already have N source, so my question is how to modify N source in MS medium for different treatments.

Ahoy,

I am working with transgenic vas::eGFP Zebrafish which I have to euthanize (ice slurry) at 29dpf. After euthanasia I am assessing the fluorescence signal of the ovaries; using fluorescence microscopy; in the whole fry, which at this stage is very small and more or less completely translucent. I am only looking at how big and how bright/fluorescent the ovaries are. I am not interested in cellular/ultracellular patterns.

Because I am working with large numbers of larvae it would be incredibly handy to be able to freeze them so that I can spread my assessment over several days instead of having to do it all in one very very frantic session.

Doe any of you have experience with freezing eGFP tissue or even better whole eGFP Zebrafish larvae?

Thank you!

Tim

ps: Yes, it will be really easy to just test it empirically but I won't have any fry ready for another 20 days so asking around is all I can do for now.

Hello everyone, i am using GFP as marker for transgenic plant confirmation but the problem is, whenever i check the GFP positive roots, there i also found too many fake/false roots (without GFP) and i need only the transgenic roots with GFP. So everytime i need to cut those negative roots that waste a lot of time. I always use antibiotics AS ( Acetosyringone ) and Timentin (TMT) (Ticarcillin disodium salt/potassium) in the MS medium when co-cultivating the explants but all in vein .

Is there any solution that i can only got positive roots with GFP?

Thanks

I'm looking for latest research articles on transcriptomics and proteomics study on transgenic sugarcane under drought stress

If yes, how to get about this problem? How to inform the consumer about the benefit and risk of "transgenic" food ?

I'm planning to do a stable transformation of BY-2 cells using Agrobacterium tumefaciens C58C1 with plasmid containing hygromycin resistance gene marker. What concentration of hygromycin I should use to select transgenic BY-2 calli?

Hello Everyone,

I did crossing of the arabidopsis (both parents were transgenic) and selected my positive F1 plants on the selection media.

I want to know how can I select homozygous plants in the future progenies?

Hi Guys!

Do anyone have experiences in making transgenic mosquitoes (A. Aedes)? We are having many difficulties in setting up the system in our lab. We couldn't find any help from neighboring labs. I'd appreciate your help.

Many thanks in advance!

W

I am very new to DREADDs and trying to find the best promotor to use for the Nucleus Accumbens, CaMK2a looks promising. Can anyone confirm that this promotor requires a transgenic animal model that expresses the CAMK2 receptor? If so, do you have a source to buy rats in the United States?

Dear,

What is the best concentration of glufosinate ammonium to select transgenic soybeans in greenhouse using spray?

I would like know the best herbicide concentration for transgenic soybean (Glycine max) selection (e.g. 35S:bar or pat genes) in greenhouse condition using glufosinate ammonium (Liberty®, BASF) spray. I've been trying to make a death curve with a national soybean cultivar using water + Silwet L-77 + 64 to 320 mg/L of glufosinate ammonium (Liberty), and spraying when the plants had the first expanded trifolium.

Thank you for feedback.

Best regards.

Marcos

What are the effects of transgenic DNA in the soil food web? What consequences, if any will these have on soil health?

In some areas where drought is persisting we observe an increase in the salinity of soils. In these areas no irrigation is possible. If you have a reference regarding this question I would greatly appreciate . Thank you!

I am working on transgenic rice. We have standarised the regeneration and transformation protocol for it. But I could see two Albino plants that are regenerated from calli. Can you please help me to short out the issue. For your kind information I want to mention that other plants regenerted with same media and hormone componets including same light condition have not shown such abnormalities. Please find the attached file.

I am a PhD student currently working with zebra finch. I think that the ability to manipulate the gene expression of specific cells within living zebra finch embryos would be useful for my research questions. Currently, our lab does not have the equipment to perform molecular approaches. For my post doc, I would like to find a lab that is currently performing genetic manipulations in birds so that I could apply these techniques to research questions in my future lab :) I would also appreciate any info about workshops or courses where I could learn about how to generate transgenic birds. Thank you!

Hi all,

I am trying to overexpress a human protein in 4T1 cells with a GFP tagged under CMV or CAG promoters. However, when we injected the cells into the 4th mammary fat pad of either NOD/SCID or NSG mice, we lost the signal after about 21 days. The tumors were about 1 cm big at the time of harvest. Have you ever got similar problems and how do you overcome it?

Thank you very much.