ASSIGNMENT # 1

Ma’aM Rida Shah

MAHAR WASEEM

(70124547)
INTRODUCTION OF CLU:
In humans, clustering is encoded by the CLU gene on chromosome 8. CLU is a
molecular chaperone responsible for aiding protein folding of secreted proteins, and its three
isoforms have been differentially implicated in pro- or ant apoptotic processes.

The CLU gene contains nine exons and expresses three isoforms alternatively-spliced at the first exon.
The encoded protein isoforms all localize to different subcellular compartments: one isoform localizes to
the nucleus; a second isoform localizes to the cytoplasm; and the third is secreted from the cell. They
also perform opposing functions: the nuclear CLU binds Ku70 to release BAX and induce apoptosis,
whereas the cytosolic and secretory isoforms inhibit apoptosis. The nuclear isoform encodes a 49 kDa
protein, while the secretory isoform, which is the main gene transcript, encodes a 75–80 kDa protein
after maturation (glycosylation, secretion, and dimerization). The mature protein is a 449-residue,
heterodimeric, disulfide-linked glycoprotein composed of two subunits of ~34-40 kDa α- and β-chains.

STEP 1:
Open the webpage of NCBI from google.
STEP 2

In the Second step, we select the nucleotide and enter the gene. Then you have to search and select
the species (Animal), molecular type (genomic DNA/ RNA), source database (GEN BANK), and
sequence type (nucleotide).

STEP 3
After opening the chosen sequence in the FASTA format selected.
How to design primer from primer 3
STEP 1
Open primer3 webpage and paste the gene sequencing in the search box

STEP 2
Click on the pick primers and you will be moved to the following page. You have to select the
left primer and click ctrl F. Now in the top of the right corner, the selected primer is shown
below in sequence in yellow color.
USE OF REVERSE COMPLEMENT.
STEP 1
Open the webpage of reverse complement from google. Paste the right primer in the box and click
submit to get the reverse complement

STEP 2
After submission of the right primer the following reverse complement was obtained.
STEP 3
Again, go on primer3 the copied sequence will paste on top of the right corner, now that
sequence appears below if it does not appear to erase some words from the left side, if it appears
then ok if it does not then erase the words from the right then it will appear below.

USE OF OLIGOCALC.
STEP 1
Go on google and open oligocalc. Now copy and paste the right primer in oligocalc then click
on calculate.
STEP 2
By clicking self-compliment the oligo self-compliment will check and the result is obtained

HOW TO DESIGN PRIMER THROUGH PRIMER FOX?

STEP 1
Search and open the Primer Fox from Google
STEP 2
After clicking on primer fox, then primer fox is opened. Paste the gene sequence in FASTA format
in the box and click submit

Step 3
After submitting the sequence, the following results were obtained.
STEP 4
Paste the right primer in the box and click submit to get the reverse complement.

STEP 5
After submitting the right primer the following reverse complement was obtained

STEP 6
The orange highlighted sequence is the reverse complement of the sequence.
 REFERENCES:
• https://pubmed.ncbi.nlm.nih.gov/11157498/
• https://www.ncbi.nlm.nih.gov/nuccore/?term=CLU
• https://bioinfo.ut.ee/primer3-0.4.0/
• https://www.bioinformatics.org/sms/rev_comp.html
• http://biotools.nubic.northwestern.edu/OligoCalc.html
• http://www.primerfox.com/