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Questions related from Md. Amzad Hossain
I have cloned a bacterial gene cluster in an expression vector (pMG36c). Each gene of the insert has its own ribosome binding site (RBS) and start codon. Reading (translation) from vector start...
10 July 2025 3,280 1 View
I have prepared competent cells of L. lactis NZ9000. But transformation not working. No colony at all in the plate. Parameters are as following: plasmid concentration: 1 uL (300ng) Electroporator-...
03 April 2024 1,213 3 View
I want to digest pET28a plasmid (5369 bp) with 2 restriction enzymes (RE)- EcoRI and XhoI for cloning. I checked the REs have single cutting site of each in the plasmid. However when I digest it...
11 June 2023 5,685 4 View
For cloning purposes, I have to double digest PCR products and plasmids 20 ug of each. Standard 50 uL restriction digestion reactions can accommodate up to 1 ug of DNA. According to some experts,...
23 May 2023 829 7 View
Hello Everyone I am trying to isolate pET28a plasmid (5.37 kbp) from E.coli strain. The electrophoresis result shows only one band that is >20 kbp. This is triplicated and same result comes all...
27 February 2023 2,366 4 View
I am planning to do Gibson assembly for the very first time. As per the recommendation of the assembly kit, 25-40 bp overlap is required between the plasmid and insert. I have designed a primer...
10 January 2023 8,359 3 View
Cloning a 6.5 kbp DNA in an expression vector failed using the heat shock method. Are there any troubleshooting points in this case?
15 November 2022 7,356 1 View
I am working with a gram positive bacteria. Please suggest a protocol for good yield.
27 October 2022 4,460 3 View
I need to transform a plasmid of 10 kbp in Gram positive competent cell (L. cremoris NZ9000). Right now, I do not have electroporation facility. Is there any successful record of heat shock...
01 January 1970 3,299 0 View
