Cloning

Clones are organisms that have identical genetic material. In other words, the sequence of bases n their DNA is exactly same. Long before the birth of Dolly the sheep, clones had bee observed in both nature and in the laboratory.When a couple has an identical twin or identical triplets, the children are clones of one another.A plant cutting can also be used to generate a clone.

Prior to 199, it was thought that cloning an entire animal could only be done with embryonic cells-cells present in the early stages of an organism’s development. In the 1950's, scientists generated entire frogs from embryonic frog cells.






After a small number of cell divisions, embryonic cells start to change into the different types of cells that form muscle, blood, liver, etc. This process is called differntiation. Although each of these cells has the same genetic material, each cell can only access the genes needed for its particular function.

Before the experiment at the roslin institute, it was thought that once cells differentiated, they could not be used to generate an entire organism, for instance, in sheep udder cells could generate other udder cells, but not an entire sheep.

The scientist of roslin institute solved this problem by growing sheep udder cells under starvation conditions, this put the cells in a state similar to embryonic cells. This is called the G0 state.

An egg cell was taken from another sheep. The nucleus (which contains the genetic material) was removed from the egg cell using fine needle. They then used electric shock to fuse one starved udder cell with one nucleus free egg cell. They made 277 of these fused cells.

Although the egg cell came from a black-faced sheep, notice that the nucleus with the genetic material came from the white-faced sheep.


The fused egg cell was then inserted into several different sheep. These surrogate mothers also black-faced.

Of the 277 fused cells, only one progressed to form a developed lamb. Dolly was born on July 5, 1996.Scientist found that dolly had same DNA as the udder cells she came from. She is a clone of these udder cells.Dolly has given birth to a lamb named Bonnie, produced the natural way.Other lambs have been born at the roslin institute through their cloning process, some carry genes that will produce usable human drugs.

A laboratory in Hawaii run by Dr.Ryuzo Yanagimachi was the second group to successfully clone an animal from an adult cell. They cloned mice using cumulus cells, a cell type found in the ovaries.

The cloning method used by the lab in Hawaii was different in two ways from the method used to clone Dolly. First, the cells used to clone the mice were not grown in culture, but instead were used immediately.

Second the nucleus was removed from the cumulus cell and then directly injected into the egg cell. This egg cell's nucleus had already been removed.

The yabagimachi lab used coat color to track genetic heritage. The cumulus cell comes from an agouti (brown) mouse, and the cell comes from a black mouse.

The egg cell now had the same genetic information as the nucleus donor mouse. The egg cell was then activated and implanted into a white host mother. On October 3, 1997 the host mouse gave birth to cumulina, named after the cumulus cells she was cloned from.


Cumulina is the same color as the mouse that donated the nucleus. The DNA fingerprinting confirmed that cumulina had the same DNA as the nucleus donor.

The scientist has taken cells from cumulina to make more clones. They have successfully made several generations of clones and all mice seem normal.Dolly the sheep died at the age of 6. Since the world said hello to Dolly, Several other animals have also been cloned.

Both Dolly and cumulina were cloned from cells in the female reproductive system; cows have also been cloned using ovary and cumulus cells with the same method that was used to clone Dolly.Pigs have been added to the cloned animal menagerie. Scientist hopes to use cloned pigs to grow organs that can be transplanted into humans.













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Alu sequence

Alu sequence is a short stretch of DNA originally characterized by the action of the Alu restriction endonuclease. Alu sequences of different kinds occur in large numbers in primate genomes. In fact, Alu sequences are the most abundant mobile elements in the human genome. They are derived from the small cytoplasmic 7SL RNA, a component of the signal recognition particle. The event, when a copy of the 7SL RNA became a precursor of the Alu sequence, took place in the genome of an ancestor of Supraprimates.



Transcript


The bulk of human genome contains many sequence that do not code for any protein. These regions of genome are sometimes referred to as "junk" Dna,and make up more than 98% pf human genome.Included in the non-coding dna are many repetitive dna sequences. Two important families of repetitive elements are LINEs(Long Interspersed Elements) and the SINEs(Short Interspersed Elements).L1 is the most prevalent LINEs, and makes up about 17% of human genome.

Alu is the most prevalent SINEs,and equals about 11% of the genome,Alu and other SINEs are "defective" transposons;they depend on the enzymes of other transposons like L1,for mobility.It is estimated that there are about 75000 L1 and 1200000 Alu elements distributed throughout the human genome. Lets take a closer look at a single copy of each


L1 ,at approximately 20 times the length of Alu,encodes all the molecular machinery needed to replicate and move within the genome.An l1 element is approximately 6000 nucleotides in length, with two untranslated regions(UTRs)and two open reading frames(ORFs).

The ORF protein products assemble into complex that enables L1 and elements such as Alu to move throughout the genome.The structure of L1 is a stark contrast with that of the tiny 300 nucleotide Alu,Alu elements have no open reading frames and so encode no proteins.


Alu elements are characterized by a sequence with two G/C rich regions: the let(L-Alu) and the right(R-Alu) monomers. An A-rich linker connects these monomers. Alu elements end with a poly-A tail.In the genome, Alu elements are immediately flanked by A=T rich sequences.

The L-Alu regions contains two specific features Box A and Box B,which are binding sites of transcription factors and RNA polymerase III.Alu elements are transcribed into RNA by these proteins until they reach a stretch of T's is the genomic DNA.

The Alu Rna can act as template for the formation of a new Alu element that can itself in a new genome location. A likely model for this process requires the enzyme, a reverse transcriptase(rt),encoded by L1.

In addition to reverse transcription,L1 rt has teh unique ability to nick DNA in a site-preferential manner. Most ofen the nick is made at the consensus sequence TTAAA.This can create a single-stranded sequences of Ts that hydrogen bonds with the poly-A tail of the Alu RNA to form a short RNA/DNA heteroduplex.

The heteroduplex than can serve as a primer for L1 rt to synthesize a complementary (c) DNA strand.

A second nick is mode on the opposite strand, a variable distance from the initial cleavage site. Then, either the L1 rt or a cellular Dna polymerase synthesizes the second DNA strand.

This results in an Alu element inserted into a novel position of the genome. The insertion also creates a direct repeat sequence on either side of the element.