Recombinant DNA technology, also known as genetic engineering, involves combining DNA from different sources to create new genetic sequences. The process includes key steps such as isolation of genetic material, cutting DNA with restriction enzymes, amplifying genes using PCR, ligating DNA molecules, and inserting recombinant DNA into host cells. Applications of this technology span agricultural modifications, medical therapies, and advancements in clinical diagnostics.

2. •The technology used for producing artificial DNA through
the combination of different genetic material(DNA) from
different sources is reffered to as recombinant DNA
TECHNOLOGY.
•Recombinant DNA technology is popularly known as
genetic engineering.
•The recombinant DNA technology emerged with the
discovery of restriction enzymes in the year 1968 by swiss
microbiologist WERNER ARBER

3. It is a series of procedures that are used to join
together (recombine) DNA segments. A
recombinant DNA is constructed from segments
of two or more different DNA molecules.
Under certain conditions ,a recombinant DNA
molecule can enter a cell and replicate
there,either on it’s own or after it has been
integrated into a chromosomes
DEFINITION:-

5. Goals of recombinant DNA technology
• To isolate and characterize a gene
•To make desired alterations in one or more
isolated genes
• To return altered genes to living cells
•Artificially synthesize new gene
• Alternating the genome of an organism
•Understanding the hereditary diseases and
their cure
•Improving human genome

6. Processof RecombinantDNA Technology
The complete process of recombinant DNA
technology includes multiple steps, maintained in a specific
sequence to generate the desired product.
Step-1. Isolation of Genetic Material.
Step-2.Cutting the gene at the recognition sites.
Step-3. Amplifying the gene copies through
Polymerase chain reaction (PCR).
Step-4. Ligation of DNA Molecules.
Step-5. Insertion of Recombinant DNA Into
Host.

7. Step-1. Isolation of Genetic Material.
The first and the initial step in Recombinant DNA
technology is to isolate the desired DNA in its pure form
i.e. free from other macromolecules.

8. Step-2.Cuttingthe geneat the recognition sites.
• DNA can be cut into large fragments by mechanical shearing.
• Restriction enzymes are the scissors of molecular genetics.
•The restriction enzymes play a major role in determining the location at
which the desired gene is inserted into the vector genome. These reactions
are called ‘restriction enzyme digestions’.

9. Restriction Enzymes —
•Primarily found in bacteria in 1960s (Werner Arber).
•A special class of sequence-specific enzyme
•site-specific-cleave DNA molecule only at specific
nucleotide sequence
•Rease recognise DNA base sequence that are
palindrome
• —Cut DNA by cleaving the phosphodiesterbond that
joins adjacent nucleotides in a DNA strand
• —Bind to, recognize, and cut DNA within specific
sequences of bases called a recognition sequence or
restriction site.

10. Step-3. Amplifyingthe genecopies throughPolymerase chainreaction (PCR).
•It is a process to amplify a single copy of DNA into
thousands to millions of copies once the proper gene of
interest has been cut using the restriction enzymes.
•Polymerase chain reaction (PCR) is a method widely used in
molecular biology to rapidly make millions to billions of
copies of a specific DNA sample,
•PCR was invented in 1983 by the American biochemist Kary
Mullis at Cetus Corporation.

11. Step-4. Ligationof DNAMolecules.
•In this step of Ligation, joining of the two pieces – a cut
fragment of DNA and the vector together with the help
of the enzyme DNA ligase.
•DNA ligase is a specific type of enzyme.it plays a role in
repairing single stranded break in duplex DNA in living
oraganisms.
Step-5. Insertion of Recombinant DNA Into Host.
•In this step, the recombinant DNA is introduced into a
recipient host cell. This process is termed
as Transformation. Once after the insertion of the
recombinant DNA into the host cell, it gets multiplied
and is expressed in the form of the manufactured
protein under optimal conditions.

14. •A vector is an area of DNA that can join another
DNA part without losing the limit for self-
replication
• Should be capable of replicating in host cell
• Should have convenient RE sites for inserting
DNA of interest
• Should have a selectable marker to indicate which
host cells received recombinant DNA molecule
• Should be small and easy to isolate

15. BACS
VECTORS
COSMID
PLASMID
EPRESSION
YACS
LAMDA
PHAGE

16. Plasmids are small, circular DNA molecules that are separate
from the rest of the chromosome.
• They replicate independently of the bacterial chromosome.
• Useful for cloning DNA inserts less that 20 kb (kilobase
pairs).
• Inserts larger than 20 kb are lost easily in the bacterial cell.

17. Lambda phage vectors are recombinant
infections, containing the phage chromosome
in addition to embedded " outside" DNA.
All in all, phage vectors can convey bigger
DNA groupings than plasmid vectors.

18. •Cosmids are hybrids of phages and plasmids that can cany
DNA fragments up to 45 kb. They can replicate like plasmids
but can be packaged like phage lambda

19. Expression vectors are vectors that carry host signals that
facilitate the transcription and translation of an inserted gene.
They are very useful for expressing eukaryotic genes in
bacteria.

20. •Yeast artificial chromosomes (YACS) are yeast vectors that have been
engineered to contain centromere, telomere, origin of replication, and a
selectable marker.
• They can carry upto 1,000 kb of DNA.
• they are useful for cloning eukaryotic genes that contain introns.

21. •Bacterial artificial chromosomes (BACS) are
bacterial plasmids derived from the F plasmid.
They are capable of carrying up to 300 kb of
DNA.

22. •Gel electrophoresis
• Cloning libraries Restriction enzyme
mapping
• PCR
• Nucleic Acid Hybridization
•DNA Microarrays

23. •DNA fragments of different sizes
can be separated by an electrical
field applied to a "gel"
•The negatively charged DNA
migrates away from the negative
electrode and to the positive
electrode.
•The smaller the fragment the
faster it migrates.

24. A DNA library is a collection of DNA fragments that have been
cloned into vectors so that researchers can identify and isolate
the DNA fragments that interest them for further study. There
are basically two kinds of libraries: genomic DNA and cDNA
libraries. Genomic DNA libraries contain large fragments of
DNA in either bacteriophages or bacterial or P1-derived
artificial chromosomes (BACs and. PACs).
cDNA libraries are made with cloned, reverse-transcribed
mRNA, and therefore lack DNA sequences corresponding to
genomic regions that are not expressed, such as introns and 5′
and 3′ noncoding regions. cDNA libraries generally contain
much smaller fragments than genomic DNA libraries, and are
usually cloned into plasmid vectors.
CLONING LIBRARIES

25. •Allows the isolation ofa specific segment of DNA
from a small DNA (or cell sample) using DNA
primers at the ends of the segment of interest.

26. •Frequently it is important to have a restriction
enzyme site map of a cloned gene for further
manipulations of the gene.
• This is accomplished by digestion of the gene
singly with several enzymes and then in
combinations.
• The fragments are subjected to gel
electrophoresis to separate the fragments by
size and the sites are deduced based on the sizes
of the fragments.

27. •A Southern allows the detection ofa
gene of interest by probing DNA
fragments that have been separated by
electrophoresis with a "labeled" probe.
• Northern Blot (probe RNA on a gel
with a DNA probe)
• Western Blot (probe proteins on a gel
with an antibody)

28. •vast majority of the protein- encoding qualities onto
a microarray chip, utilizing innovation in light of the
DNA silicon chip industry.
• The chip can be utilized to hybridize to cell RNA,
and measure the statement rates of a substantial
number of qualities in a cell.

29. •Agriculture: growing crops of your choice (GM food),
pesticide resistant crops, fruits with attractive colors, all being
grown in artificial conditions
• pharmacology: artificial insulin production, drug delivery to
target sites
•Clinical diagnosis – ELISA is an example where the
application of recombinant
• Medicine: gene therapy, antiviral therapy, vaccination,
synthesizing clotting factors
• Other uses:fluorescent fishes, glowing plants etc
NEW APPROACHES
•Used in treatment of organ failure:
ie; XENOTRANSPLANTATION
•Used in design,development,isolation of proteins

30. •https://www.slideshare.net/nasira55/recombinant-dna-technology-
47715143
•https://byjus.com/biology/recombinant-dna-technology/
•https://en.wikipedia.org/wiki/RecombinantDNA
•https://www.medicinenet.com/script/main/art.asp?articlekey=5247
•https://www.slideshare.net/pragatiRandive/recombinant-dna-
technology-and-its-applications
•https://www.slideshare.net/MahendraMahi28/recombinant-dna-
technology-142600446
•https://www.britanica.com/science/recombinant-DNA-technology