I am working with a lot of bacterial isolates from the environment. I've already sequenced them and many are from different species. I try best to keep them pure and not mixed-up during my experiment, so that I know which one is which. Some isolates are easily to track because of morphological difference such as pigment or colony appearance. However, I get stuck sometimes and I am not sure some isolates I am working on are still the same isolates as previous. I have no good ways to track them and ensure they are the original isolates during my project. Do you have any suggestions?
I hope you are following proper culture maintenance practice, while subculturing some population may change its genetic make up due to mutation, it is better to keep a lyphophilized stock or glycerol stocks.
For traceability purpose, we can follow a bar coding system. But a proper labeling with self coding system is more than enough, until unless when your lab have been used by many people, then bar coding system will be a better one.
from the very beginning, try to save your culture in duplicate under the same condition, whenever need them, take only one to do your experiments while keep the other one intact. handling the bacteria follow the wel estabolised protocols to minimize potential contamination.
You'd need to have replicates of your culture, stored properly at 4 °C and/or freezed at -80 °C. Lyophilization is also a possibility. Also you should characterize your cultures by molecular techniques (MSP-PCR or RAPD or other ready-repeatable technique) in order to be able to compare. If not, you'll lose track of them (if you are working with mixed cultures and want to know how are the culture evolving).
if you will be running a series of experiments, grow up a batch of each strain and store multiple vials of each at -80. Go back to the freezer to use the original stock each time you start.
Perhaps during your work you must use several strains in parallel and in successive processes, in this case I recommend to design small DNA primer, 16sRNA region targeting (FISH), it's not very expensive. You can even identify two strains present in an same experiment if each primer has a different fluorochrome. The limitation is that the microscope must be equipped with appropriate filters. Here you can see the description of the method.
http://www.scirp.org/journal/PaperInformation.aspx?PaperID=35674#.U9pDuoB5PgQ
Initially obtaining pure cultures is of prime importance. Further, try to maintain at least 2 sets of glycerol stocks out of the pure culture obtained, of which one should never be taken out (Freezing and thawing repeatedly kills bacteria).
Colony morphology, as of colony size and pigmentation may vary after stocking the bacteria.
you have to preserve and store slant culture with use Teflon coated screw cap tube differentiate color indication labeling system and each an every time subculture.
Make number of long storage stocks (eg. glycerol, Lyophilization stocks) and performed Amplified Ribosomal DNA Restriction Analysis (ARDRA) initially, so you can match ARDRA profile of culture, whenever you want.
You can lyophilise it. If you dont have such facility, kindly store them any national microbial depository, it will be cost effective and you can get your culture back whenever you want.
- Badges
- Science topic
- Similar topics
- Biological Science
- Microbiology
- Microorganisms
- Bacteria