Unexpected colony growth on Rifampicin plates with E.coli DH5Alpha and K-12?

last month 0 8K Report

I am a Bachelor student working on an Experiment. I am currently working with E.coli DH5 Alpha, K-12 and Agrobacterium tumefaciens strain EHA105.

When i plated they strains onLB Agar containing kanamycin,Hydromycin and rifampicin, none of the strains grew, which was expected.

However, when i plated the strains EHA105, K-12 and DH5alpha on LB Agar containing only rifampicin, EHA105 grew as expected, but after 3-4 days i observed a large circular colony on the DH5 alpha and K-12 which shouldn’t be resistent against rifampicin. During the first 2 days there was no visible growth.

My question is: could this late appearing colony be due to cross-contamination with EHA105, or could it be a spontaneous rif-resistent mutant or maybe a fungal contamination ? Does anyone who has encountered such problems a solution for me ?

Badges
Science topic

More David Emeh's questions See All

Is it possible to conjugate an NHS ester to the N-terminus of a peptide ON RESIN?

So i have a few peptides (synthesized by SPPS and not cleaved from resin) and I want to add a maleimide label. We have NHS-PEG-Maleimide and from literature it is evident that the free amine group...

18 July 2024 4,075 3 View

Hot to do MTT assay data analysis?

I have repeated an MTT assay three times using different batches of the same cell line. Each time, I tested the same treatment (different medium) and seeded the same number of cells. Additionally,...

08 July 2024 1,900 1 View

What is wrong with Site-Directed Mutagenesis?

I am following the NEB protocol to perform SDM with Q5 and NEB HiFi Assembly. I am making a two base pair change. First picture is my Primers designed with NEBaseChanger. Second Picture is my...

24 June 2024 6,079 4 View

Does anyone have suggestions for this esterification?

I have a very polar alcohol (soluble in MeOH, H2O, partially in heated DMSO). And a long chain fatty acid that I would like to couple. I can't find a common solvent for them. I can form the...

24 June 2024 9,319 0 View

How do you add multiple nodes or cutting planes in Abaqus?

Hi, In a previous question I asked how to effectively model springs in Abaqus. However, I figured out the feature under Assembly -> springs/dashspots -> node to node Springs. I have a cylinder...

20 June 2024 892 3 View

How do you add radial spring elements in Abaqus?

Hi, I am trying to complete a buckling analysis on a tube with an external pressure. This solves itself easily by adding radial constraint on the internal edge of the tube, one axial constraint...

17 June 2024 8,165 7 View

Given that Mesozoic placentals were largely arboreal (with fingers), do you know of any studies that illustrate the finger-to-hoof transition?

I'm interested in the earliest days of placental terrestrial locomotion in large herbivorous taxa. Phenacodus seems to be transitional in this regard. Any others?

06 June 2024 5,182 2 View

How reliable is ninhydrin test in solid-phase peptide synthesis?

Hey everyone! I'm doing peptide synthesis on Rink amide resin and sometimes I need to add amino acids with secondary amines or beta amino acids. Normally I synthesize 35 to 40 mer peptides. As a...

21 May 2024 9,035 0 View

How does one generate a normalized TCSPC curve from actual photon counts versus time?

Sometimes TCSPC curves are analyzed using the actual photon counts, and sometimes they are normalized with the signal running from 0 to 1, or even from 0 to some value less than 1. What quantity...

20 May 2024 4,277 0 View

Problem when running Spin-Orbit Coupling using CASTEP. What is nlpot_prepare_precon_ks?

I am currently using CASTEP version 22.11, and I encounter the following error when running calculations with spin-orbit coupling: Error with dsytrf in nlpot_prepare_precon_ks Current trace...

19 May 2024 4,236 5 View

What is the best way to drop a plasmid?

Please address the best way to drop a plasmid. Background: I have a "bait" plasmid resistant to kanamycin and a "prey" plasmid resistant to carbenicillin. After many rounds of streaking on...

25 July 2024 2,532 3 View

Pink bacterial colonies?

I am cloning an overexpression plasmid with my protein of interest tagged with mScarlet. After transforming my ligated product into DH5α bacteria and plating on LB agar, I noticed colonies with a...

22 July 2024 5,953 6 View

My rosetta strain has chloramp resistance and i am transforming my plasmid with amp resistance but not getting colonies on chloramp/amp LB plates?

I have done the same procedure with BL21 and pLysE but i got colonies but multiple times on rosetta i did but not getting. can anyone explain what would have gone wrong?

12 July 2024 9,904 1 View

Has anyone calculated the effective concentration at 50% (EC50) value of different fungicides for the fungus Tilletia horrida?

How can we measure their colony diameter?

12 July 2024 7,027 0 View

I have constucted a plasmid vector from puc19 with no antibiotic and lacz gene. The problem is screening for transformed cells. Any suggestions?

Get too many colonies, which are mostly non-transformants when tested by colony PCR

11 July 2024 6,775 3 View

I am not getting proper colonies while ligating one of fragments in my vector the colonies seems to be normal but in light it seems lysed any reason?

I am working on cloning of a gene fragment inside my vector everything we tried to do correctly like making insert and vector with utmost care but after ligation the morphology of colonies seems...

11 July 2024 9,010 3 View

Can someone provide any tips for cell quantification on Fiji, ImageJ?

Hi. I have brightfield images of stem cells. I need to count the total number of cells/image. Some cells are individual and some cells are in colonies. Using average cross-sectional area of...

05 July 2024 4,197 1 View

Cloning restrction digestion needed before sequencing?

Hello, recently i am an issue swith cloning where I am using the subclone method to insert my gene length is 1.5kb and my vector(pMAL-c6T) is 5kb I got the colonies which i screened through the...

04 July 2024 7,977 2 View

Why I am unable to get the expected insert at the cloning site?

I am trying to clone a gRNA (23 bases) into the Crispr cas9 pDirect23A vector by using restriction enzyme cloning. I have performed a complete experiment, starting with plasmid isolation,...

02 July 2024 5,448 2 View

Why my transformed colonies do not carry my insert?

I digest my plasmid with KpnI-NotI. I cut the expected 4kb band from the gel, purified it and then I did ligation [1:3 and 1:5 vector:insert ratio] and transformation. I found 18 transformed...

30 June 2024 3,976 0 View