SU Course: Methods in Molecular Life Sciences Part – 2

Today I will talk about the wet lab aspect of the Practicals of the methods course.

2. Lab Practicals

The main aim of the lab exercise was to generate mutant strains of the Saccharomyces cerevisiae and look for the mutations in genes involved in Lysine production  pathway using Illumina sequencing.

1. Experimentation and Data generation – Wet lab

a. Yeast Culturing

We started by culturing yeast in plates from the culture flasks by using different OD to get a good growth that can be used for further experimentation. Saccharomyces cerevisiae or commonly known as Yeast takes about 2 days for good growth. Thus, after streaking on the YAPD (rich media) we let them grow for about 2 days.

b. Clonal Purification

The yeast cells were then clonally purified on nitrogen-poor media that contained high levels of α-Aminoadipate and lysine. This is an intermediate from the Lysine production pathway. However, downstream synthesis product derived from this intermediate is shown to cause the growth inhibition on nitrogen poor media. Thus, if cells do not have the 3 important lysine genes (LYS2, LYS5, LYS14) then the conversion of α-Aminoadipate to the toxic intermediate does not take place, allowing the cells to grow.

Selecting clones for plating and obtaining genetically purified (identical) colonies

Thus, we clonally select cells that have mutation in these Lysine genes or so called auxotrophic for Lysine.

c. Validation of auxotrophy

To verify that the cells thus selected actually have the mutations in the lysine genes, they are replica plated on Lysine free media to see for no growth as opposed to lysine rich media.

The third column shows a mutant strain, as it has no growth in Lysine deficient media

d. Complementation test

Plate containing the auxotrophic mutants was crossed with the tester strains (strains that are known to have mutations in LYS2, LYS5 and LYS14 genes) by replica plating.  This test is used to determine experimentally the specific gene in which each mutant has a mutation. This observation is used to obtain preliminary information if it is worth sequencing the sample and where to expect the mutation while analysing sequencing data. Good samples were selected and used to perform Illumina sequencing.

Replica plating for complementation test (crossing tester strains with the mutants from the velvet to the plate)

SU Course: Methods in Molecular Life Sciences Part-1

The next course in the Masters is of Methods in Molecular Life Sciences. This course is also a two part course like the Bioinformatics course.

The two parts are :

1. Theory lectures

2. Lab practicals

1. Experimentation and data generation – Wet lab

2. Data analysis – Dry lab

1. Theory lectures

There are 10 theory lectures each 2 and 1/2 hours long. The classes start at 10 in the morning until lunch time. Each class focuses on different techniques and methods in molecular biology – wet lab. All these methods are taught by experts in the field, who use these techniques in their labs on daily basis. The classes are conducted by faculties from the MBW department at Stockholm University and Scilife lab Fellows.

The main topics covered in the course are Protein-protein interaction. This class is complementary to the course on Lipids and bio-membranes. The next lecture is introduction to various Protein-nucleic acid interaction, where they show how various Transcription factors which are proteins recognize the nucleic acids for their function.

Following this, we had two lectures on Yeast as a model organism and molecular tools available for use in this organism. This also formed basis to the lab exercises that we performed in the afternoon, after the lectures. Continuing the discussion on the tools, we were introduced to the more tools that can be used on various model systems and in-vitro mutagenesis, ethical concerns and some interesting examples of the use of some of them in the on-going research in Dr. Claudia Kutter’s lab.

Dr. Claudia Kutter’s lecture on gene modification

While mentioning model systems, we were also about microbial growth systems, and a recent advancement in the technique of single cell imaging. Moving to higher Eukaryotic organisms we learned about Drosophila melanogaster, were we saw the first manual drawing of the polytene chromosome. Further, we learned the epigenetic basis of DNA modification and how it can be used to create desired mutations chromosome wide.

Manual drawing of the polytene chromosomes from Fruit fly

After the Easter break of about 2 weeks, we had the most interesting lecture of the course on the advancements in the next generation sequencing techniques by Dr. Vicent Pelechano. In this, we were fascinated to see how the same samples can be pre-processed in multiple ways to get different kind of information (TSS, TTS, splicing information, snap shot of the genes currently transcribed, and many more…) from it, by employing the same Illumina sequencing at the last step. Finally, we had a introductory lecture to the data analysis explaining its importance in the age of Big Data.

One such seuencing method – TIFseq, developed in Dr. Vicent Pelechano’s lab      Source: Nature journals

Appointment for Biometrics

Once you have submitted your application for the Residential permit, and paid the fees, the next step is to book an appointment for submitting your biometrics.

Swedish residential permit is in the form of a card. This card contains your biometrics information. This includes fingerprints, signature and photo. In order to submit this information with the migration agency, you need to go to the nearest Swedish embassy. You can find the information about the nearest embassy on the website here.

The option to select the embassy will appear after the payment section in your application. Select the embassy that is nearest to you from the above link and state that in the given column. Then you will have an option to select the slot convenient to you from the available slots. Note that the submission of biometrics does not affect the decision on your case. You can alternatively wait and book the appointment for biometrics after you receive the decision. However, I recommend to not wait that long as the slots might fill be full at a later time.

While at the embassy, you have an option to get you card delivered to your home address or to collect it from the embassy. To get the residential permit card delivered to your home address there is a form to be filled that is provided by the staff at the embassy. The residential permit generally takes about 2-3 weeks to arrive at your home address after the decision has been made and the biometrics have been submitted. You can find the information of the cards dispatched from the Sweden every week on the Sweden-abroad website in the waybills.

In case of queries you can write to the Swedish migration agency from their contact page. Alternatively you can call them on the given number. They generally answer in short time.

 

Applying for Residential Permit

The Visa for Sweden is called as he Swedish Residential Permit. If you are a non-EU (non-European Union) country, then you will probably need a residential permit.

Application for Residential Permit or RP is relatively easy. The application is made online using the Swedish Migration agency’s website. The Swedish Migration agency is called Migrationsverket. You can find the link to the website here. The website is in Swedish, however, you can view this in different languages, there is an option at the top of the page to change languages. I recommend using this option, sometimes google translate might not be accurate and can also cause trouble loading some options.

On the main webpage, there is an option to apply for a new application. From the drop-down list select the option of ‘Permission to study’. This new page has the detailed information of everything. It is best to spend some time and read all the information before starting the application.

I am writing a checklist that can be used as a reference before starting the application. The requirements for the document is very specific and needs to be followed strictly. A deviation from the required format can cause delay in the decision making process. For further details please check the website.

Checklist before starting the application:

  1. Scanned copy of Passport. It has to be scanned properly, including all the corners.
  2. Download the admit letter (Notification of selections) from the universityadmissions.se. This is the proof of acceptance. Further, you can also upload the welcome letter from your home department. (It can take some time for the students of the joint masters programmes to get this).
  3. FAS+ insurance. This is provided to all the international fee paying students by KTH. This is to be uploaded as health insurance certificate.
  4. Bank statement of the sum required to for the application from a single holder bank account. The account must be in the name of the applicant only. (Statements from the joint holder’s account are not accepted).
  5. Receipt of the Tution fee payment.
Bank statement requirements for application

Once you have the documents ready, click on the start application. You have to select the option of ‘Private Individual’ and login option of using the ‘Username and Password’. Create a new account using an email ID. Make sure to have an email ID that’s accessible an in routine use. You will receive all the important information on this ID.

Once you have the account ready, login using the same and follow the instructions. The application period will be of 13 months. You will have to extend the permit after a year from Sweden.

Application login options (Use the one highlighted with the blue arrow)

The information that you have filled in your application is saved for a short period of time (generally 2-3 days). You will be able to see this information when you log out from your application. However, the information can still be found later, but it is best to submit in the given period just to avoid the need to fill everything again from start.

The whole process is very straight forward. The best way to get a quick decision is to submit all the documents in the required format and according to the given specifications. However, if there is any kind of problems, then you can contact the migration agency by email, they respond very quickly. Further, after submitting the application, if they find something missing or incomplete, then the migration agency gets back to you with the requirements of extra information and the details on how to submit them.

Next step is to book an appointment for biometrics. I will write about this soon.  So stay tuned!

KTH tution fee payment

I have been admitted, but what to do next? How to confirm my admission with KTH? How to apply for Swedish visa? How to get housing, etc etc. Do not worry about these questions. I will be writing a series of posts to guide you through the process.

Once you have received an offer, first step would be to confirm your position. For an international tution fee paying student, this can be done by paying the tution fees for the first semester. You will receive the invoice with the tution fees for the first semester. The due date to pay the fees and confirm your admission is 31st May, 2019.

For the EU students, generally there is no specific process to be followed as such to confirm your admission.

Credit card

The easiest way to pay the tution fees is by a credit card.

For this, open the link mentioned in the email that says ‘pay.kth.se’. This will redirect you to a payment portal as shown below. There it will ask for Invoice/OCR number and customer ID. These are mentioned on the top right corner of the invoice. Fill these and follow the instructions on the screen.

Fee payment portal

Bank Transfer

However, it is relatively difficult to have credit cards with such high credit limit, so most people opt for bank transfer option.

Most of the banks have a form that needs to be filled that is specific to the bank. So the best way is to go to the branch of your bank and enquire about money transfer or wire transfer. The bank employees should then guide you through the process.

Invoice. Yellow color highlights the important details

However, one of the most important thing is to mention the customer number and the Invoice number, on the bank forms of wire transfer. These numbers are mentioned on the top right side of the Invoice. The forms have a blank space where you can mention these. Sometimes there is no specific provision on the forms to mention anything, in that case you should ask the staff about this and ensure that these numbers are mentioned during the wire transfer. This is how KTH will identify your payment and confirm your admission.

Bank transfer generally take 3-4 working days, so it is a good idea, to start the process early and have some buffer time of at about a week before the due date.

Hope this was helpful. If you have any relevant doubts then feel free to drop in an email.

My next blog will be on ‘How to apply for the Swedish Visa or Swedish residential permit’. So stay tuned!