Azucares Reductores

Introduction
14 i/ham

to recombinant-DNA

technology1’2

L Carroll
Recombinant-DNA in virtually brief to used every identify aspect genes, of this exposition technology of the is to to provide isolate the is now biological an DNA that make up a mammalian sciences. located on either DNA strand outline of be oriented in opposite directions gene The cornchromosome. and on the thusindividual a chromosome. Genes genes One may he may gene

ABSTRAC’l’
rnonlv The the terest. short gating used purpose approaches to amplify introduction along and

the gene if necessary. and to the principles ofseparating with a description these large of an genes. mm

of incodes for a single polypeptide: to clone genes. isA illustrated in Figure 2. very large genes (mRNA) (gene transcription) toNiur

flow synthesis

of genetic information of messenger RNA by genetic ofthe RNA elements gene. These polymerase core secontrol.

is controlled portion enzyme

is provided.

approach J (liii

cloning

propa- that usually lie 5’ to the ,)ro,flok’r elements serve to the quences These

protein-coding to focus the

I 993:58(suppl):249S-58S.

KEY
RNA chain

WORDS
polvrnerase.reaction. \east

Plasmid,
cloning. artificial

bacteriophage. reverse
restriction endonuclease. chromosome

transcriptase. polymerase

transcription start point. Many promoters share that are essential for effective transcription core sequences (bo.ves) include the thymine-adenine-thy(TATA) box, and box. the cytosine-adenine-adenine-thythe guanine-cytosine (GC) box. upstream(5’) whereasstart point for in exons.

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mine-adenine mine (CAAT)

The mRNA regions

Introduction

CAAT and GC boxes the TATA box seems synthesis. The genes

are located further to control the actual ofeukaryotes are

arranged

of the gene represented in the mature mRNA. Exons are sepaThe overall goal of recombinant-DNA technology is toidenrated by intervening sequences or introns. which are spliced from tify, isolate. manipulate. and re-express genes from a given host RNA template during RNA processing. Other eu( I -9). Some of the practical goals of such cut-and-paste tech-the primary karyotic-mRNA processing events include the addition of a nology is to 1) develop a basic understanding of the function methylated guanine residueconnected by a 5-5’ triphosphate and regulation of known gene products. 2) identify new genes to the first nucleotide ofthe mRNA (cap site) as well as whose protein products have not been isolated (reverse genetics). linkage the addition of a stretch of polyadenylic acid residues at the 3’ 3) correct endogenous genetic defects (eg. sickle cell anemia). 4) terminus. The mRNA is translated intoprotein with a methiexpress foreign genes in disease-susceptible hosts (eg. diseaseserving as a translation start site. Proteins also have polarity resistance genes in agricultural crops). and 5) manufacture large onine with an unpaired amino group (NH2) at one end and a free quantities ofa protein product for widespread use (eg. antibodies carhoxyl group at the other. The amino-terminal part of thein tobacco plants). Any discussion ofthis methodology should begin ecule by with itself. hydrogen a description DNA bonds ofthe between unique oftwo their features antiparallel nitrogenous-base ofthe DNA side mol-protein bound is encoded by the 5’ end ofthe mRNA. is composed strands

chains. Restriction endonucleases Genetic information is provided by the purine and pvrimidine The firstbreakthrough in recombinant bases. which are linked to a sugar phosphate backbone (Fig). 1 of enzymes capable This structural support is a series of deoxyribose residues linked the identification reproducible fragments. b phosphodiester bonds, which are created by covalently joining into discrete a hydroxvl group at the 3-carbon residue with a phosphate group adjacent sugar group. Thus DNA unpaired 5’...