PRACTICLE 1.

Method of searching biological data by Accession number

Accession number
A unique number given to an entry in biological databases that serves as its permanent identifier.

Procedure
1. First of all open NCBI site.
2. Open gquery databases page
3. Add your accession number in search bar and search.

4. Select gene from the database page

5. Following information is found;
 6. Select genomic sequence below and get its FASTA sequence.
DNA sequence of gene is given below;
GCCATGTATATTTTTATTATTTCAGTAATAACATGCGAGTGAGAAAATAGTATAAATATTAGTGGAAAAAG
ACAGACTTCCTTAATGGATATAAACATCAAAATTCAATAATTGCGATAGAGTATCAATCAAGATCTTACAT
ATAAAAAAAGATTCATCCCCCAAAGAACGGAACACTCACTCTTCTAAAACGATGGGAGAGATAGCAAC
AGAGTTCACGCCGGTTATGGATGATGATGATGACCGGTGCGTTGTACCGGAGGTAGAGCTCACTGTACC
CAAGACCGACGATTCAACTTTACCGGTTTTAACATTTCGGATGTGGGTTCTTGGTATAGGCGCGTGCATT
GTACTCTCCTTTATCAACCAGTTTTTCTGGTACAGAACGATGCCGTTGAGTATCACCGGAATCTCCGCTC
AGATCGCCGTCGTGCCGCTTGGTCATCTCATGGCGAGAGTGCTTCCGACAAAAAGATTCTTGGAAGGTA
CGAGGTTTCAGTTTACCTTAAACCCGGGTGCCTTCAACGTCAAGGAACATGTTCTGATAACTATCTTTGC
AAATTCCGGCGCCGGCTCTGTTTACGCTACTCACATACTTAGTGCTATTAAGCTTTACTACAAACGTTCTC
TCCTTTCCTCCCTGCATTTCTCGTCATGATCACCACTCAGGTACTCTCACTTCACTTTATCCTCTGTTTCTT
GTTTTGCAGTTTATTGAACCTTGAAATCGTTTCTTGTGTTTAGATTCTTGGATTTGGTTGGGCTGGTCTCT
TCAGGAAACACCTTGTTGAGCCTGGTGAAATGTGGTGGCCAAGCAATCTTGTTCAAGTCTCTCTCTTTG
GTGCGTTACACGAGAAAGAAAAGAAATCAAGAGGAGGCATGAGTAGAACTCAATTCTTCTTAATCGTC
CTCGTAGCTAGCTTTGCTTACTACATCTTTCCTGGTTATCTTTTCACCATGTTAACATCAATCTCATGGGTC
TGTTGGCTCAATCCCAAATCAATTCTGGTAAACCAACTCGGTTCAGGTGAACATGGTCTTGGCATTGGTT
CCATTGGTTTTGATTGGGTCACTATTAGTGCTTATCTTGGAAGCCCTTTAGCTAGTCCTTTGTTCGCTTCC
GTCAATGTAGCCATTGGTTTCGTGCTGGTTATGTATATCGTCACGCCTGTTTGTTACTGGCTTAACATTTAT
GATGCCAAGACCTTCCCTATCTTCTCTAGTCAGCTCTTCATGGGGAATGGCTCACGTTATGATGTTTTGAG
CATTATTGATAGCAAATTCCATCTTGACCGTGTGGTTTACTCTAGGACCGGGTCCATCAATATGAGCACCT
TCTTTGCTGTTACTTATGGTCTCGGGTTTGCTACATTGTCTGCCACCATTGTCCATGTTTTAGTCTTCAAC
GGAAGGTACATATATTCAATGATGTTTACCAACACTTCTTTTTGTATCTTGGTCCCAAACTCAGACTAATC
TTTATGTCTATACAGTGACTTGTGGAAGCAGACAAGAGGGGCGTTTCAGAAGAATAAGAAATGGATATA
CACACGAGAATCATGAAGAAAAACTACAGGGAAGTTCCATTGTGGTGGTTTTTGGTAATCCTTTTACTG
AATATCGCGCTTATCATGTTCATCTCCGTGCACTACAACGCTACTGTGCAGCTTCCTTGGTGGGGAGTGTT
ACTTGCTTGTGCCATTGCCATCAGTTTCACTCCTTTGATCGGTGTCATAGCTGCCACCACTAATCAGGCA
CCGGGATTGAATATCATAACAGAGTATGTTATCGGGTATATCTACCCGGAACGTCCTGTTGCAAACATGTG
TTTCAAGGTCTATGGATACATTAGTATGACTCAAGCGCTAACTTTCATTTCCGACTTCAAGCTTGGTCATT
ACATGAAGATCCCGCCTAGATCCATGTTCATGGCTCAGGTATGCATGTTATCAATTGGTCTTGAGACAGAT
GTCAATAAAACTAAAACATAGCTAATCAAACTGTAACAAGTGCAGGTGGCTGGGACACTTGTGGCGGTG
GTTGTGTACACAGGAACAGCCTGGTGGTTAATGGAAGAGATTCCTCATCTTTGTGATACATCCTTGCTTC
CTTCAGACAGCCAATGGACTTGTCCCATGGACCGCGTTTTCTTTGACGCCTCTGTGATTTGGGGACTAGT
GGGACCACGCAGGGTGTTTGGAGACCTTGGAGAATACTCAAATGTCAACTGGTTCTTCCTCGTTGGAGC
CATAGCTCCACTCTTAGTTTGGCTAGCCACAAAGATGTTTCCGGCTCAGACATGGATCGCCAAGATTCAT
ATCCCTGTCCTTGTGGGAGCCACCGCAATGATGCCACCAGCAACAGCAGTTAACTTCACTAGTTGGCTC
ATCGTCGCATTCATCTTTGGCCACTTCATCTTCAAGTACAGAAGAGTGTGGTGGACCAAGTATAACTATG
TGTTGTCTGGTGGTTTAGATGCTGGCTCTGCTTTTATGACCATACTTCTGTTTCTAGCACTCGGACGAAA
AGGAATTGAAGTGCAGTGGTGGGAAATTCCGGCGACCGAGATACATGTCCCTTAGCTTCTTGTCCAACT
GCTAAAGGTGTTGTCGTCAAAGGCTGTCCCGTCTTCTAGCTCATGAGTCATGAGACAAAAGCATATACC
AAGTGCTTTGTACAGAACACACAGCAAACTGATAATTGTAATTCTCGGAAATGTCTTTACTCTCCAAGAA
CTATATCTTTTGTGATAGCTTTAAAACTAGGTACTATCCTGTTCTTTGACTGAACACAAATGCTAGTGATT
CTATATGTGGTTCCTTAAAATGATTTTAATGCATGCTTCAGAAAGA

PRACTICLE 2.
Primer Designing
Primers are short fragments of DNA that are used to recognize and amplify a particular region of
DNA. It is about 18-20 bp long.
There are different designs used for primer designing some of the following are;
1. Primer 3
2. Auto prime
3. Prime bank etc.
Most commonly used tool is Primer 3.

Procedure
1. First download the DNA sequence.
2. Paste your DNA sequence in Primer 3 software

3. Adjust primer size up to 18-20 base pairs.

4. Adjust melting temperature (Tm) from 50-65 c.
5. Adjust product size between100-200.
6. Select pick primer and we got forward and reverse primers.

Checking of Primer Specificity

 1. Open primer blast tool.
2. Paste your DNA sequence and forward (left) and reverse (right) sequences in primer
parameters.

3. Select genome data for all organism in primer pair specificity checking parameters
data bases
4. Get primers

PRACTICAL 3
BLAST
BLAST is a tool that is used to search relevant sequences in a data base. It finds regions of
similarity between biological bases.
Principle
It compares nucleotide sequences and protein sequences to the data base and calculate statistical
significance of their matches.
Procedure
1. Open NCBI and go for BLAST tool.

2. Then click on nucleotide BLAST.

3. Click blastn for nucleotide BLAST and blastp for protein sequence match.
4. Now paste your sequence ( FASTA format) on search place.

5. Click BLAST and you get the following results.

PRACTICLE 4.
Building of phylogenetic tree by Matching of Multiple sequences
It is matching of more than two sequences using BLAST and other tools (Mega 7).
Multiple matching gives us the following benefits.

 It predicts conserved domains

 It also gives phylogenetic analysis

Procedure
1. First of all use blast tool and get a sequence match of your unknown sequence.
2. Now open Mega 7 tool for multiple match.
3. Click on align and select edit or build alignment.
4. Select a new alignment and choose for DNA or Protein.

5. Paste your sequences in blue spot and toggle it on 100%. Conserved matches in all
sequences is highlighted.
6. Now go to Data option and click phylogenetic analysis.

7. Now minimize the page and go to phylogeny in tool bar.

8. Select construct maximum likelihood tree.
9. Click ok and compute.
10. You get the final tree.

11. Its random tree form is