Status and Overview of Biotechnology in ... - ACS Publications

Chapter 1 Status a n d O v e r v i e w o f Biotechnology in Agricultural Chemistry Homer M. LeBaron

Downloaded by 80.82.77.83 on May 24, 2018 | https://pubs.acs.org Publication Date: March 18, 1987 | doi: 10.1021/bk-1987-0334.ch001

Ciba-Geigy Corporation, Greensboro, NC 27419

Biotechnology has become the scientific buzzword of our age. One definition of "buzz" is "a confused murmur or flurry of activity." To a large extent this is a fitting explanation of what is going on under the diverse umbrella of biotechnology. No doubt the new techniques and tools, and the rapidly expanding knowledge of biology, genetics and biochemistry justify the enthusiasm, popularity, and expectations. Finally, mankind can experience the exuberance of having the power to "create life" with a new dimension. We are learning that this power s t i l l has limitations in spite of its great potential. Within this chapter are described some of the exciting scope and breadth of potential applications and products of biotechnology in the areas of crop and microbial production. This treatise is by no means complete or exhaustive, but it will hopefully provide a reasonable list to enlighten the layman and to motivate the expert. In addition, many of the basic tools and techniques used in biotechnology are outlined and described briefly. Some projections on the probabilities or possibilities of success, both short-term and long-term, as well as patent problems and some views on environmental and health concerns are also discussed.

E x p e c t a t i o n s and L i m i t a t i o n s J u s t p r i o r t o the symposium on " A p p l i c a t i o n s o f B i o t e c h n o l o g y t o A g r i c u l t u r a l Chemistry" h e l d a t the ACS m e e t i n g i n Chicago, September 9-12, 1985, John K. Crum, E x e c u t i v e D i r e c t o r o f ACS, asked the p e r t i n e n t q u e s t i o n , "Does the American Chemical S o c i e t y have a r o l e i n b i o t e c h n o l o g y ? " The symposium and t h i s subsequent p u b l i c a t i o n s e r v e as a s t r o n g c o n f i r m a t i o n t h a t t h e f u t u r e o f b i o t e c h n o l o g y i s not o n l y c o m p a t i b l e w i t h , but dependent on c h e m i s t r y as w e l l as many o t h e r areas o f s c i e n c e . 0097-6156/87/0334-0001 $06.00/0 © 1987 American Chemical Society

LeBaron et al.; Biotechnology in Agricultural Chemistry ACS Symposium Series; American Chemical Society: Washington, DC, 1987.


2

BIOTECHNOLOGY IN AGRICULTURAL CHEMISTRY

The e s s e n t i a l t e c h n o l o g i c a l b r e a k t h r o u g h t h a t l e d t o t h e c u r r e n t e x p e c t a t i o n s and p o t e n t i a l f o r b i o t e c h n o l o g y and, t h e r e by, the b i r t h o f g e n e t i c e n g i n e e r i n g , took p l a c e i n 1973 when S t a n l e y Cohen o f S t a n f o r d U n i v e r s i t y and H e r b e r t Boyer o f t h e U n i v e r s i t y o f C a l i f o r n i a a t San F r a n c i s c o found a s i m p l e way t o combine DNA from two d i f f e r e n t organisms and then t o c l o n e i d e n t i c a l c o p i e s o f these recombined DNA m o l e c u l e s i n b a c t e r i a . D u r i n g t h e past t e n y e a r s , we have been inundated w i t h c l a i m s as t o how g e n e t i c e n g i n e e r i n g w i l l r e v o l u t i o n i z e our lives. These r e v o l u t i o n a r y changes were t o r e s u l t from t h e development o f new wonder d r u g s , n o v e l v a r i e t i e s o f crop p l a n t s and r a d i c a l l y d i f f e r e n t methods o f r a i s i n g e c o n o m i c a l l y i m p o r t a n t farm a n i m a l s . R e c e n t l y , a prominent s c i e n t i s t gave an exaggerated n o t i c e t h a t most o f those i n v o l v e d i n c u r r e n t a g r i c u l t u r a l c h e m i s t r y , p l a n t and a n i m a l b r e e d i n g and o t h e r r e l a t e d t e c h n o l o g y w i l l need t o r e t i r e e a r l y o r be r e t r a i n e d s i n c e t h e r e w i l l be no need f o r p e s t i c i d e s and many o t h e r o f our p r e s e n t t o o l s (J_). T h i s statement i s j u s t as s e r i o u s an e x a g g e r a t i o n as those made by some s c i e n t i s t s i n the 1950's as the many s p e c t a c u l a r h e r b i c i d e s were d i s c o v e r e d , t h a t we c o u l d now do away w i t h t h e c u l t i v a t o r , hoe, and m e c h a n i c a l weed c o n t r o l . While i t i s t r u e t h a t t h e new t o o l s c r e a t e d by b i o t e c h nology w i l l r e p l a c e some o f our p r e s e n t methods, they w i l l , i n most c a s e s , p r o v i d e a d d i t i o n a l means o f d o i n g our jobs b e t t e r . For a wide v a r i e t y o f r e a s o n s , t h e i r i n t r o d u c t i o n and impact w i l l be a slow and g r a d u a l p r o c e s s . Reasonable p r e d i c t i o n s f o r i t s impact on w o r l d food p r o d u c t i o n i n d i c a t e t h a t a 5 t o 10 p e r c e n t i n c r e a s e may r e s u l t from b i o t e c h n o l o g y i n t h e next q u a r t e r c e n t u r y , and t h a t p l a n t g e n e t i c s might add 5 t o 20 b i l l i o n d o l l a r s a year t o t h e v a l u e o f major crops by t h e 1990's. The f i r s t g e n e t i c a l l y e n g i n e e r e d c r o p p l a n t s w i l l l i k e l y come on the market sometime between 1986 and 1990, w i t h o n l y a few p r o d u c t s at f i r s t . By t h e year 2000, some p r e d i c t t h a t t h e new t e c h n o l o g y may dominate a g r i c u l t u r e (J_, 2_> 3) · P l a n t s c i e n t i s t s have a c h i e v e d i m p o r t a n t and s i g n i f i c a n t r e s u l t s i n t h e past decade. However, t h e achievements t o date i n p l a n t s c i e n c e do n o t b e g i n t o approach t h o s e o f s c i e n t i s t s c l o n i n g u s e f u l mammalian genes i n v a r i o u s c e l l s and m i c r o organisms. The main reasons f o r t h i s marked c o n t r a s t a r e t h a t the number o f s c i e n t i s t s i n t e r e s t e d i n p l a n t p h y s i o l o g y and g e n e t i c s a r e few, and t h e f i n a n c i a l s u p p o r t f o r t h e s e areas o f b i o t e c h n o l o g y i s s m a l l , compared t o t h e r e s o u r c e s a v a i l a b l e i n mammalian o r m i c r o b i a l g e n e t i c s . More i m p o r t a n t l y , t h e base l i n e o f knowledge a v a i l a b l e i n p l a n t g e n e t i c s i s much l e s s than t h a t a v a i l a b l e i n m i c r o b i a l , o r even a n i m a l c e l l g e n e t i c s . Even i f the p l a n t genes c o d i n g f o r such i m p o r t a n t economic f a c t o r s as drought t o l e r a n c e , d i s e a s e r e s i s t a n c e , p e s t i c i d e t o l e r a n c e , or i n c r e a s e d y i e l d were c h a r a c t e r i z e d and i d e n t i f i e d , which i s n o t y e t t h e case, the a b i l i t y t o m a n i p u l a t e and t r a n s f e r these genes at w i l l t o most p l a n t s i s v e r y l i m i t e d . I t can be s t a t e d w i t h c o n v i c t i o n t h a t , i n g e n e r a l , s u f f i c i e n t b a s i c knowledge necessary t o r o u t i n e l y and e f f e c t i v e l y e n g i n e e r new and improved v a r i e t i e s o f major c r o p p l a n t s v i a recombinant DNA does not e x i s t today.



LEBARON

Status and Overview

There i s no doubt t h a t the p r o d u c t s of b i o t e c h n o l o g y and g e n e t i c e n g i n e e r i n g w i l l have, i f they haven't a l r e a d y had, a major impact on our c u r r e n t j o b s and what many of us are d o i n g . Any a r e a of s c i e n c e or r e s e a r c h tends t o grow and then d e c l i n e . B i o t e c h n o l o g y i s growing today as o t h e r d i s c i p l i n e s are d e c r e a s i n g i n importance or p o p u l a r i t y . A l t h o u g h most o f us w i l l not need to be c o m p l e t e l y r e t r a i n e d , except by c h o i c e , we w i l l c e r t a i n l y need t o be f l e x i b l e , w i l l i n g to change and t o develop new s k i l l s , and always keep l e a r n i n g . T h i s book, h o p e f u l l y , w i l l s e r v e as p a r t of t h i s process f o r some. L i k e many f i e l d s of s c i e n c e , the u l t i m a t e b e n e f i t and a p p l i c a t i o n of our d i s c o v e r i e s must o f t e n be found i n o t h e r d i s c i p l i n e s , or i n i n t e r a c t i o n s and c o m b i n a t i o n s of d i s c i p l i n e s . We must r e c o g n i z e t h a t b i o t e c h n o l o g y , as c h e m i s t r y , i s a t o o l , a means to produce t o o l s , or a means t o an end, t h a t serves i n o t h e r f i e l d s . We should a l s o a p p r e c i a t e t h a t fame, f o r t u n e , i n f l u e n c e and p r e s t i g e are u s u a l l y f r a g i l e ; as e a s i l y and q u i c k l y l o s t as g a i n e d . Those of us who have l i v e d through such c y c l e s know t h a t b i o t e c h n o l o g y w i l l not always be the magic word and s c i e n t i f i c a t t r a c t i o n t h a t i t i s today. That i s not because i t i s o v e r e s t i m a t e d or doesn't deserve the p o s i t i o n and esteem i t e n j o y s , but o n l y t h a t i t i s new and the r e a l i t i e s , l i m i t a t i o n s and dependence upon o t h e r s k i l l s are not yet f u l l y r e c o g n i z e d . The a t t e n t i o n o f the p u b l i c , p o l i t i c i a n s , and i n v e s t o r s w i l l soon be d i v e r t e d t o o t h e r p r i o r i t i e s , long b e f o r e the f u l l impact and b e n e f i t s of b i o t e c h n o l o g y have been d i s c o v e r e d or d e v e l o p e d . I f the p o s s i b i l i t i e s i n h e r e n t i n b i o t e c h n o l o g y seem e n d l e s s , man's t r u s t i n s c i e n c e i s n o t , and we have seen many s i g n s o f rebellion. I t i s u n f o r t u n a t e t h a t even now, i n i t s i n f a n t s t a g e , the p r o d u c t s and e a r l y commercial a p p l i c a t i o n s o f b i o t e c h n o l o g y a r e b e i n g t h r e a t e n e d by s k e p t i c s who a l r e a d y f e a r t h a t i t s r i s k s w i l l be g r e a t e r than i t s b e n e f i t s . As Dr. Robert Kaufman of Monsanto Company r e c e n t l y s t a t e d , " I f i t ' s g o i n g to c o s t $2.5 m i l l i o n e v e r y time we make a minor m o d i f i c a t i o n i n a gene, then t h i s f i e l d i s dead" ( 4 ) . Most o f us would agree t h a t the f e a r s and o b j e c t i o n s o f the c r i t i c s are o f t e n unfounded or e x a g g e r a t e d , but few of us would d i s a g r e e w i t h t h e i r i n s i s t e n c e t h a t the burden o f r e a s o n a b l e p r o o f f o r e n v i r o n m e n t a l and h e a l t h e f f e c t s s h o u l d be on those promoting t h e i r commercial use. Nor s h o u l d we c l a i m t h a t e x a g g e r a t i o n s about b i o t e c h n o l o g y a r e the e x c l u s i v e p r o p e r t y of the n o n - s c i e n t i s t s and uninformed c r i t i c s . We have been exposed t o about e q u a l doses of s c i e n c e f i c t i o n both from the promoter of the m e r i t s and f u t u r e p o t e n t i a l o f our s c i e n c e as from the hypochondriac who imagines t h a t manmade monsters are l u r k i n g i n the t e s t tubes w a i t i n g t o t a k e over the w o r l d . One o f the most ardent and v o c a l c r i t i c s o f b i o t e c h n o l o g y , Jeremy R i f k i n , warns t h a t i t s p r o d u c t s w i l l d e s t r o y mankind 05). In a more r a t i o n a l p l e a , Marc Lappé u r g i n g more s o c i a l and e t h i c a l r e s p o n s i b i l i t y i n the a p p l i c a t i o n s of b i o t e c h n o l o g y , s t a t e d "Recombinant DNA r e a l l y i s the m i l l e n n i u m i n b i o l o g y , j u s t as t h e d i s c o v e r y of n u c l e a r f i s s i o n was the m i l l e n n i u m i n p h y s i c s . The power t o do good or i l l from both t e c h n i q u e s i s v i r t u a l l y l i m i t l e s s . We need not exaggerate our c o n c e r n about what a few mad s c i e n t i s t s may i n a d v e r t e n t l y c r e a t e i n a l a b o r a t o r y o r



4


r e l e a s e i n t o t h e environment, but we must h o l d a c c o u n t a b l e what whole i n d u s t r i e s o r governments w i l l choose t o c r e a t e w i t h t h i s newfound power" ( 6 ) . I t i s q u i t e p o s s i b l e t h a t t h e g l o r y years o f b i o t e c h n o l o g y w i l l be s h o r t e r than f o r most p r e v i o u s scientific disciplines. Containment o r c o n t r o l a f t e r r e l e a s e , e s p e c i a l l y o f e n g i n e e r e d m i c r o o r g a n i s m s , seems t o be the g r e a t e s t problem. A b i l i t y t o p r o p e r l y m o n i t o r t h e p r o d u c t s o f b i o t e c h n o l o g y ( e . g . m i c r o b e s ) and c o n t i n g e n c y p l a n s i n case o f containment problems a r e a l s o o f c o n c e r n . While b i o t e c h n o l o g y has t h e p o t e n t i a l t o be a much needed and major i n d u s t r y f o r t h e American economy, i t i s f a c i n g a r e g u l a t o r y o b s t a c l e course t h a t has f r e q u e n t l y bogged i t down i n l e n g t h y and l a b y r i n t h i n e a p p r o v a l p r o c e s s e s . Even worse, once f i e l d t e s t s o r a p p l i c a t i o n s o f b i o t e c h p r o d u c t s have been approved, the d e c i s i o n has sometimes been t h r e a t e n e d by, or fought o u t , i n t h e c o u r t s . A s u p p o r t i v e , r e a s o n a b l e , and c o o r d i n a t e d r e g u l a t o r y environment, a l o n g w i t h a c o n t i n u i n g e d u c a t i o n a l e f f o r t aimed at t h e p u b l i c and government, must be e s t a b l i s h e d b e f o r e b i o t e c h n o l o g y can a c h i e v e s u c c e s s , e s p e c i a l l y i n the United States ( 7 ) . P o t e n t i a l Products o f Biotechnology I t i s not a p p r o p r i a t e o r p o s s i b l e t o enumerate h e r e a complete l i s t o f p o t e n t i a l p r o d u c t s or b e n e f i t s from b i o t e c h n o l o g y . However, some o f t h e major t r a i t s o r p r o d u c t s p r e s e n t l y b e i n g developed o r c o n s i d e r e d i n e n g i n e e r e d p l a n t s and microorganisms a r e l i s t e d i n Table 1 and T a b l e 2, r e s p e c t i v e l y . While some g e n e t i c c h a r a c t e r i s t i c s c o u l d be u s e f u l i n both p l a n t s and m i c r o o r g a n i s m s , and b a c t e r i a l genes w i l l o f t e n be used i n e n g i n e e r i n g o f crop p l a n t s , the products or t r a i t s are l i s t e d a c c o r d i n g t o t h e most l i k e l y end u s e . Many o f t h e s e t r a i t s a r e not d i s c u s s e d i n d e t a i l i n t h i s p u b l i c a t i o n . We have l i m i t e d t h e scope o f t h i s book and symposium p r i m a r i l y t o c r o p , s o i l and environmental a p p l i c a t i o n s , with very l i t t l e a t t e n t i o n given t o a n i m a l and human uses o f b i o t e c h n o l o g y . Some o f t h e e f f e c t s o f b i o t e c h n o l o g y on a g r i c u l t u r a l c h e m i s t r y w i l l be i n d i r e c t c o m p e t i t i o n w i t h o r i n o p p o s i t i o n t o t h e use o f c h e m i c a l s , such as t h e development o f pest r e s i s t a n t crops t a k i n g the place o f present p e s t i c i d e s . Other a p p l i c a t i o n s w i l l enhance o r a l l o w new uses f o r a g r i c u l t u r a l c h e m i c a l s , such as: (1) the t r a n s f e r o f h e r b i c i d e r e s i s t a n c e i n t o o t h e r w i s e s u s c e p t i b l e c r o p s , ( 2 ) the use o f g e n e t i c a l l y a l t e r e d m i c r o organisms t o enhance the d e g r a d a t i o n o f t o x i c o r p e r s i s t e n t c h e m i c a l s i n t h e environment, and ( 3 ) t h e use o f e n g i n e e r e d m i c r o o r g a n i s m s i n t h e s y n t h e s i s and c o m m e r c i a l p r o d u c t i o n o f s p e c i f i c isomers or c h e m i c a l s f o r use i n a g r i c u l t u r e . I t has long been p r e d i c t e d t h a t c r o p p l a n t s r e d e s i g n e d t o be r e s i s t a n t t o h e r b i c i d e s w i l l be t h e f i r s t p r o d u c t s o f b i o t e c h n o l o g y o f p l a n t s (j$). There a r e a number o f reasons why t h e g e n e t i c e n g i n e e r i n g f o r h e r b i c i d e r e s i s t a n c e i s so a t t r a c t i v e , i n c l u d i n g the f o l l o w i n g :


1.

LEBARON

Status and Overview Table 1 P o t e n t i a l P l a n t Improvements and P r o d u c t s From B i o t e c h n o l o g y


1.

2.

3.

H e r b i c i d e r e s i s t a n c e i n c r o p s . T h i s i s one of the s i m p l e s t , most p o p u l a r , and p r o b a b l y w i l l be among the f i r s t e n g i neered t r a i t s t o be of commercial a p p l i c a t i o n . I t has a l s o become a common development method and a model system from which t o generate background e x p e r i e n c e i n t r a n s f o r m i n g plants. I t c o u l d be one o f the most p r o f i t a b l e approaches f o r i n d u s t r i e s t h a t can s e l l the h e r b i c i d e - r e s i s t a n t crop seed as w e l l as more h e r b i c i d e . A wide range of crops r e s i s t a n t t o many h e r b i c i d e s ( e . g . , t r i a z i n e s , g l y p h o s a t e , paraquat, i m i d a z o l i n o n e s , s u l f o n y l u r e a s , e t c . ) i s under development o r i n r e s e a r c h . In the case of some of these e f f o r t s ( e . g . , a t r a z i n e - r e s i s t a n t s o y b e a n s ) , the o b j e c t i v e may be more f o r the purpose of a v o i d i n g c r o p p h y t o t o x i c i t y o r s o i l c a r r y - o v e r i n j u r y from p r e v i o u s a p p l i c a t i o n s than f o r d i r e c t use of the h e r b i c i d e on the r e s i s t a n t c r o p . There are l i k e l y more than 100 companies or l a b o r a t o r i e s i n the U.S. where b i o t e c h n o l o g y r e s e a r c h i s underway on herbicide resistance. Seed or s t o r a g e p r o t e i n s a l t e r e d or i n c r e a s e d by the i n s e r t i o n o f genes t h a t a l t e r the p r o d u c t i o n of c e r t a i n amino a c i d s , enhancing the q u a l i t y and n u t r i t i o n a l v a l u e of seeds or g r a i n s . For example, much of our c o r n c r o p i s fed to a n i m a l s , but i s d e f i c i e n t i n p r o t e i n c o n t a i n i n g e s s e n t i a l amino a c i d s , e s p e c i a l l y l y s i n e . A number of h i g h l y s i n e mutants have been developed u s i n g c l a s s i c a l g e n e t i c m a n i p u l a t i o n . The most n o t a b l e i s the opaque-2 v a r i e t y of c o r n . More r e c e n t l y , h i g h l y s i n e r i c e has been d e v e l o p e d . In the case o f both r i c e and c o r n , the improvement i n l y s i n e c o n t e n t has l e d t o s i g n i f i c a n t l o s s e s i n y i e l d when the c r o p i s grown under f i e l d c o n d i t i o n s . I n the case of h i g h l y s i n e c o r n , o t h e r agronomic d i f f i c u l t i e s have been encountered, such as h a r v e s t i n g , s t o r a g e , and proc e s s i n g problems, due i n l a r g e p a r t t o the s o f t n e s s of the c o r n k e r n e l i n the new v a r i e t y . The premium p r i c e p a i d f o r the h i g h l y s i n e corn i s not h i g h enough to o f f s e t the 10 t o 20 percent lower y i e l d which the farmer o b t a i n s , and h i g h l y s i n e c o r n today i s not a v i a b l e economic crop ( 1 5 ) . The seeds of legumes and c e r e a l g r a i n s p r o v i d e humans w i t h an e s t i m a t e d 70 p e r c e n t of t h e i r d i e t a r y p r o t e i n r e q u i r e m e n t s , but the p r o t e i n s i n t h e s e seeds are a l s o d e f i c i e n t i n c e r t a i n e s s e n t i a l amino a c i d s . Researchers have examined the p o s s i b i l i t y of g e n e t i c a l l y e n g i n e e r i n g t h e genes t h a t code f o r these p r o t e i n s t o a l t e r t h e i r amino a c i d c o m p o s i t i o n , but they are complex, m u l t i g e n e f a m i l i e s . A l s o , i f we modify a s i n g l e amino a c i d , i t might a f f e c t the secondary and t e r t i a r y s t r u c t u r e o f the p r o t e i n , which a f f e c t s how i t i s f o l d e d and d e p o s i t e d as a s t o r a g e p r o t e i n (2). D i s e a s e r e s i s t a n c e i n c r o p p l a n t s , such as e n g i n e e r i n g p l a n t s t o t u r n on d e f e n s i v e mechanisms.


5

BIOTECHNOLOGY IN AGRICULTURAL CHEMISTRY 4.

5. 6. 7.


8. 9.

10.

11.

12.

13. 14. 15.

I n s e c t r e s i s t a n c e i n c r o p p l a n t s , by t r a n s f e r r i n g genes t h a t enable p l a n t s t o make t h e i r own i n s e c t i c i d e s (BT t o x i n - see T a b l e 2, item #3). Drought r e s i s t a n c e i n crop p l a n t s . T o l e r a n c e t o f l o o d i n g , excess m o i s t u r e and poor s o i l aeration. T o l e r a n c e t o s a l t , s a l i n i t y or t o x i c metals i n crop p l a n t s , which would a l l o w crops t o be i r r i g a t e d w i t h sea o r b r a c k i s h water, or t o be grown i n v a s t areas not now s u i t a b l e because of h i g h a l k a l i or s a l t c o n t e n t . T o l e r a n c e t o temperature extremes i n crop p l a n t s . I n c r e a s e the c y s t e i n e and m e t h i o n i n e amino a c i d s ( h i g h sulfur protein) in a l f a l f a . T h i s i s b e i n g attempted by A u s t r a l i a n s c i e n t i s t s by i n s e r t i n g pea genes (p-albumin-1) i n t o a l f a l f a , a n t i c i p a t i n g enhanced wool p r o d u c t i o n i n sheep ( 16). The m o d i f i c a t i o n of l e a f or f o r a g e p r o t e i n s i s p r o b a b l y more f e a s i b l e than s t o r a g e p r o t e i n s , but t h e r e are l i k e l y sharp upper l i m i t s ( 2 ) . More e f f i c i e n t a b s o r p t i o n and u t i l i z a t i o n o f f e r t i l i z e r s . The i n c r e a s e d uptake o f phosphorous i n e a r l y c r o p d e v e l o p ment has been p r o m i s i n g . E x t e n d i n g and i n c r e a s i n g the a b i l i t y of n i t r o g e n f i x a t i o n t o major c r o p s . T h i s c o u l d g r e a t l y reduce our dependency on n i t r o g e n f e r t i l i z e r s and the c o s t of crop p r o d u c t i o n . I t has been r e s e a r c h e d e s p e c i a l l y i n c e r e a l g r a i n s , r i c e and c o r n , but has not appeared p r o m i s i n g due t o the l a r g e number o f genes i n v o l v e d ( 1 7 ) , and the f a c t t h a t b i o l o g i c a l n i t r o g e n f i x a t i o n consumes enormous amounts o f energy and would have t o be f u e l e d , f o r the most p a r t , by metabolism o f c a r b o h y d r a t e s from the p l a n t i t s e l f (_2, 15). I n c r e a s e the p h o t o s y n t h e t i c e f f i c i e n c y of c r o p p l a n t s . Much r e s e a r c h has been done on the genes t h a t code f o r the enzyme ribulose-1,5-bisphosphate carboxylase-oxygenase (Rubisco), which i s p r o b a b l y the most abundant p r o t e i n i n the w o r l d and the key c a t a l y s t i n p h o t o s y n t h e s i s . Rubisco c o n s i s t s of e i g h t l a r g e p r o t e i n s u b u n i t s encoded by genes i n c h l o r o p l a s t s and e i g h t s m a l l s u b u n i t s encoded by genes i n t h e n u c l e u s . A l t h o u g h no v e c t o r systems c u r r e n t l y e x i s t t o a l t e r genes i n the c h l o r o p l a s t s , r e s e a r c h e r s are g e n e t i c a l l y e n g i n e e r i n g the genes t h a t encode the s m a l l s u b u n i t s . By i n c r e a s i n g the e f f i c i e n c y w i t h w h i c h R u b i s c o f i x e s atmospheric carbon d i o x i d e , r e s e a r c h e r s hope t o produce p l a n t s t h a t w i l l grow f a s t e r . Development of s h o r t e r season v a r i e t i e s . M o d i f y growth stages or h a b i t s of c r o p p l a n t s f o r improved harvestability. Improvement i n t a s t e or t e x t u r e o f v e g e t a b l e s and f r u i t s , such as b e t t e r t a s t i n g c e l e r y s t a l k s and tomatoes w i t h h i g h e r s o l i d s content ( 1 3 ) . Somaclonal v a r i a t i o n s f o r s e l e c t i o n of d e s i r a b l e v a r i a n t s , and somatic embryos ( a r t i f i c i a l seeds) have been e s p e c i a l l y u s e f u l i n g e n e t i c e n g i n e e r i n g o f improved v e g e t a b l e s . S e v e r a l of these new c r o p v a r i e t i e s w i l l be on the market w i t h i n 2 t o 4 y e a r s .


LEBARON

16.

17.

18.

Status and Overview

I n c r e a s e d crop y i e l d or f a s t e r g r o w t h . These t r a i t s w i l l be v e r y d i f f i c u l t f o r the g e n e t i c e n g i n e e r t o m a n i p u l a t e d i r e c t l y s i n c e they p r o b a b l y i n v o l v e many genes and i n t e r actions. I t w i l l be e s s e n t i a l t o determine how many genes and o t h e r c o m p l i c a t i n g or l i m i t i n g f a c t o r s a r e i n v o l v e d i n t h i s and o t h e r t r a i t s b e f o r e b e g i n n i n g a g e n e t i c e n g i n e e r i n g project. New f l o w e r s , h o r t i c u l t u r a l c r o p s , and ornamental crops w i t h improved t r a i t s , such as f a s t e r b l o o m i n g , g r e a t e r f l o w e r i n g , etc. Develop c r o p s t h a t produce and e x c r e t e a l l e l o p a t h i c c h e m i c a l s o r p r o p e r t i e s t o b e t t e r compete w i t h weeds o r insects.


Table 2 P o t e n t i a l M i c r o b i a l and Other Improvements Products From B i o t e c h n o l o g y 1.

2.

3.

4.

and

I c e - n u c l e a t i o n a c t i v e b a c t e r i a . T h i s was the f i r s t m i c r o b i a l product o f b i o t e c h n o l o g y f o r which a p p r o v a l of f i e l d t e s t s was r e q u e s t e d . T h i s has r e s u l t e d i n a long c o u r t b a t t l e w i t h no f i n a l c o n c l u s i o n t o d a t e . A f t e r 241 experiments, e x p e n d i t u r e s of $750,000, and f i v e man-years of r e s e a r c h , Advanced G e n e t i c S c i e n c e s has not yet c o n v i n c e d r e g u l a t o r s and the p u b l i c t h a t t h e i r m o d i f i e d Pseudomonas f l u o r e s c e n s ( F r o s t b a n ) i s s a f e . The l a t e s t news on the s i m i l a r ice-minus b a c t e r i a (P. s y r i n g a e ) b e i n g developed by Dr. Steven Lindow and the U n i v e r s i t y o f C a l i f o r n i a , B e r k e l e y , i n d i c a t e s t h a t f i e l d t e s t s w i t h t h i s microbe w i l l a l s o be d e l a y e d u n t i l 1987. Improve the s t r a i n s of n i t r o g e n - f i x i n g b a c t e r i a (Rhizobium) o r the mechanism by which they f i x n i t r o g e n i n p l a n t s . T h i s has a t t r a c t e d new a t t e n t i o n s i n c e i t was found t h a t many genes are r e s p o n s i b l e and a h i g h energy l o s s t a k e s p l a c e when p l a n t s (e.g. legumes) f i x n i t r o g e n , making i t v i r t u a l l y i m p o s s i b l e t o t r a n s f e r t h i s t r a i t i n p l a n t s at the p r e s e n t time. Develop a more t o x i c or cheaper s t r a i n of B a c i l l u s t h u r i n g i e n s i s (BT), or expand and improve d e l i v e r y and a p p l i c a t i o n systems. Many companies are d e v e l o p i n g new t e c h n o l o g i e s i n v o l v i n g g e n e t i c m a n i p u l a t i o n o f BT. Rohm and Haas and s e v e r a l o t h e r companies have i n s e r t e d t o x i n p r o d u c i n g BT p r o t e i n s i n t o chromosomes o f t o b a c c o , attempting to provide p r o t e c t i o n against c e r t a i n insects throughout the l i f e c y c l e of the c r o p . Monsanto has i m p l a n t e d t o x i c BT p r o t e i n i n s t r a i n s o f Pseudomonas f l u o r e s c e n s , which they then use t o coat c r o p seeds ( e . g . c o r n ) t o p r o t e c t the c r o p r o o t s from i n s e c t (rootworm) a t t a c k . Mycogen uses a s i m i l a r a p p r o a c h , but k i l l s the b a c t e r i a by heat treatment b e f o r e a p p l i c a t i o n ( 1 7 ) . G e n e t i c e n g i n e e r i n g of o t h e r m i c r o b i a l and v i r a l insecticides.


BIOTECHNOLOGY IN AGRICULTURAL


5.

6.

7.

8.

9. 10.

11.

12. 13.

CHEMISTRY

Development o f microorganisms o r p r o d u c t i o n o f enzymes f o r biodégradation o f p e s t i c i d e s and o t h e r t o x i c wastes. This i s a l a r g e and d i v e r s e p o t e n t i a l t h a t has l o n g been o f i n t e r e s t t o m i c r o b i o l o g i s t s and o t h e r s i n v o l v e d i n h a n d l i n g and p r e v e n t i n g e n v i r o n m e n t a l c o n t a m i n a t i o n from p e s t i c i d e s p i l l s , w a s t e s , c o n t a i n e r c l e a n - u p , p l a n t e f f l u e n t s , on-farm s p r a y r i n s a t e s , e t c . G e n e t i c e n g i n e e r i n g has g r e a t l y enhanced t h e i n t e r e s t and o p p o r t u n i t i e s i n t h e s e a r e a s , w h i l e t h e r e c e n t awareness and l o n g - t e r m c o n c e r n about groundwater c o n t a m i n a t i o n make i t u r g e n t and i m p e r a t i v e t h a t t h i s t e c h n o l o g y be d e v e l o p e d . I t has been e s t i m a t e d t h a t Superfund c l e a n - u p s w i l l c o s t up t o $100 b i l l i o n over t h e next f i v e y e a r s ( 1 8 ) . Most o f t h e c l e a n - u p w i l l merely r e s u l t i n moving t h e problem from one p l a c e t o a n o t h e r . The o n l y t o t a l l y s a t i s f a c t o r y and permanent s o l u t i o n i s t o c l e a n up t h e h a z a r d i n s i t u o r where i t o c c u r s . B i o t e c h n o l o g y o f f e r s the best and cheapest p o t e n t i a l l o n g - t e r m f o r h a n d l i n g most o f t h e s e problems ( 1 9 ) . G e n e t i c a l l y engineered p l a n t pathogens w i l l p r o v i d e a d d i t i o n a l t o o l s t o study p a t h o g e n i c i t y (e.g. mutation a n a l y s i s , stage o f i n f e c t i o n , host s p e c i f i c i t y and r e s i s t a n c e , e t c . ) , l e a d i n g t o improved c r o p r e s i s t a n c e and disease control. Use o f monoclonal a n t i b o d i e s t o d e t e c t and q u a n t i f y s p e c i f i c pathogens, d i f f e r e n c e s between v i r u l e n t and n o n - v i r u l e n t pathogens, and pathogen r e s i s t a n c e t o p e s t i c i d e s ( f u n g i c i d e s ) , and t o prevent d i s e a s e by c o n f e r r i n g p a s s i v e immunity t o an i n f e c t i o u s agent. Use o f monoclonal a n t i b o d i e s and immunoassays f o r both f i e l d and l a b o r a t o r y d e t e r m i n a t i o n s ( q u a l i t a t i v e and q u a n t i t a t i v e ) of many e n v i r o n m e n t a l l y and c h e m i c a l l y s t a b l e p e s t i c i d e s , m e t a b o l i t e s and o t h e r a g r i c u l t u r a l c h e m i c a l s . Many companies and l a b o r a t o r i e s a r e a c t i v e l y d e v e l o p i n g and u s i n g t h i s t e c h n o l o g y . W h i l e i t w i l l not c o m p l e t e l y r e p l a c e GC, HPLC and o t h e r c h e m i c a l assay methods f o r many a n a l y t i c a l r e q u i r e m e n t s , immunoassay methods a r e p o t e n t i a l l y much f a s t e r , e a s i e r and cheaper, once t h e y a r e i n p l a c e and o p e r a t i n g , and they can be even more s e n s i t i v e and s p e c i f i c (20). They can u s u a l l y p r o v i d e q u a l i t a t i v e r e s u l t s w i t h i n one t o two m i n u t e s , and q u a n t i t a t i v e r e s u l t s w i t h i n f i v e minutes t o a few h o u r s , and w i l l have a wide range o f u s e s . They w i l l d e t e c t p e s t i c i d e i n u r i n e , b l o o d , a i r , groundwater, f o o d , p l a n t s and s o i l . Development o f m y c o h e r b i c i d e s o r s i t e - s p e c i f i c b e n e f i c i a l p l a n t pathogens. Development o f microbes t h a t a r e a b l e t o p r o t e c t crop p l a n t s from adverse e n v i r o n m e n t a l f a c t o r s , such as s o i l s a l i n i t y , a c i d i t y and t o x i c m e t a l s . Development o f m i c r o b i a l and e n z y m a t i c systems f o r p r o d u c t i o n o f n a t u r a l product c h e m i s t r y f o r a g r i c u l t u r e , i n c l u d i n g p e s t i c i d e s , secondary m e t a b o l i t e s (e.g. b i a l a p h o s , w i t h h e r b i c i d a l a c t i v i t y ) , o p t i c a l isomers, s p e c i a l t y chemicals, e t c . Development o f microbes f o r improved f e r m e n t a t i o n and u t i l i z a t i o n o f a g r i c u l t u r a l biomass. Use o f p l a n t c e l l c u l t u r e systems t o produce secondary metabolites or chemical substances.



1.

LEBARON

Status and Overview

1) I t i s r e l a t i v e l y s i m p l e . The b i o a s s a y f o r t h e r e s i s t a n t t r a i t can be f a s t , r o u t i n e , and the r e s i s t a n t c e l l s are e a s i l y s e p a r a t e d from s u s c e p t i b l e c e l l s or p l a n t s . 2) H e r b i c i d e r e s i s t a n c e or t o l e r a n c e has o c c u r r e d f a i r l y r e a d i l y i n n a t u r e ( e s p e c i a l l y t o t r i a z i n e s ) ( 8 , 9^. 3) Mechanisms o f h e r b i c i d e a c t i o n and r e s i s t a n c e have been w e l l s t u d i e d and they have o f t e n been dependent on a s i n g l e gene. 4) H e r b i c i d e r e s i s t a n c e i n weeds can be u s e f u l , and has a l r e a d y been c o m m e r c i a l l y t r a n s f e r r e d t o c r o p s by c o n v e n t i o n a l breeding. 5) C o n s i d e r a b l y more money i s spent on h e r b i c i d e s f o r weed c o n t r o l than f o r a l l o t h e r t y p e s o f p e s t i c i d e s combined. 6) In s p i t e of the g r e a t e f f o r t s and c o s t s f o r weed c o n t r o l , i t i s e s t i m a t e d t h a t weeds s t i l l c o s t more than $20 b i l l i o n i n l o s t crop p r o d u c t i o n per year i n the U.S. (9_> 10). 7) The c o s t of g e n e t i c a l l y e n g i n e e r i n g a c r o p and e x t e n d i n g t h e use of e s t a b l i s h e d h e r b i c i d e s i s much cheaper than d e v e l o p ment c o s t s f o r a new h e r b i c i d e , assuming the t e c h n o l o g y i s a v a i l a b l e and r e g u l a t o r y c o s t s are not p r o h i b i t i v e . 8) Many of our best and most e c o n o m i c a l h e r b i c i d e s cause s o i l c a r r y o v e r , f o l i a r i n j u r y , or o t h e r s e l e c t i v i t y problems which can be remedied by d e v e l o p i n g r e s i s t a n t c r o p s . 9) The b a s i c r e s e a r c h on h e r b i c i d e r e s i s t a n c e p r o v i d e s important new knowledge on p l a n t p h y s i o l o g y , b i o c h e m i s t r y , and g e n e t i c e n g i n e e r i n g . 10) H e r b i c i d e r e s i s t a n c e i n c r o p s o f f e r s f l e x i b i l i t y i n weed c o n t r o l methods which i s v e r y i m p o r t a n t , e s p e c i a l l y under c o n s e r v a t i o n t i l l a g e and w i t h m i n o r c r o p s . 11) H e r b i c i d e r e s i s t a n t crops c o u l d s e r v e t o extend the l i f e o f p a t e n t s or expand use of h e r b i c i d e s i n some c h e m i c a l markets. In o r d e r t o f i n d and m a n i p u l a t e the genes t h a t code f o r r e s i s t a n c e t o h e r b i c i d e s , i t i s e s s e n t i a l t h a t we understand the mechanisms o f a c t i o n of h e r b i c i d e s i n p l a n t c e l l s . It is f o r t u n a t e t h a t most h e r b i c i d e s i n t e r a c t w i t h enzymes and o t h e r p r o t e i n s i n m e t a b o l i c pathways s p e c i f i c t o p l a n t s , such as amino a c i d s y n t h e s i s and p h o t o s y n t h e s i s . T h e r e f o r e , they are g e n e r a l l y n o n - t o x i c t o a n i m a l s , and g e n e t i c e n g i n e e r i n g f o r these t r a i t s i s s p e c i f i c for plants. F o r example, g l y p h o s a t e i n h i b i t s the enzyme, EPSP ( 5 - e n o l p y r u v y l s h i k i m a t e 3-phosphate) s y n t h a s e , t h a t c a t a l y z e s a s t e p i n the s y n t h e s i s of the a r o m a t i c amino a c i d s . S i m i l a r l y , both the i m i d a z o l i n o n e s and s u l f o n y l u r e a s i n h i b i t a c e t o l a c t a t e synthase (ALS), the enzyme t h a t c a t a l y z e s the f i r s t s t e p i n the f o r m a t i o n o f branched-chain amino a c i d s ( 1 1 ) . T r i a z i n e h e r b i c i d e s act by b i n d i n g t o a s p e c i f i c p r o t e i n i n the t h y l a k o i d membranes o f the c h l o r o p l a s t s , p r e v e n t i n g the f l o w of e l e c t r o n s and i n h i b i t i n g p h o t o s y n t h e s i s ( 1 2 ) . U s i n g d i f f e r e n t g e n e t i c e n g i n e e r i n g s t r a t e g i e s , Monsanto and Calgene s c i e n t i s t s have developed g l y p h o s a t e r e s i s t a n t p l a n t s . Monsanto developed g l y p h o s a t e r e s i s t a n t p e t u n i a s , tobacco and tomatoes by e n g i n e e r i n g o v e r p r o d u c t i o n o f EPSP synthase i n these


9


10


plants. Calgene has moved a gene f o r a mutant EPSP enzyme, which i s not i n h i b i t e d by g l y p h o s a t e i n t o t o b a c c o , c o t t o n , tomato and poplars. In a d d i t i o n , M o l e c u l a r G e n e t i c s has d e v e l o p e d c o r n p l a n t s r e s i s t a n t t o American Cyanamid's i m i d a z o l i n o n e h e r b i c i d e , and DuPont r e s e a r c h e r s have i d e n t i f i e d mutant ALS genes, which c o n f e r s u l f o n y l u r e a r e s i s t a n c e , but t h e y have not as yet r e p o r t e d g e n e t i c a l l y e n g i n e e r e d r e s i s t a n t p l a n t s (2^, _U, 13). W h i l e i t i s d i f f i c u l t t o p r e d i c t w h i c h p r o d u c t s and a p p l i c a t i o n s of b i o t e c h n o l o g y w i l l succeed or be of most i m p o r t a n c e , i t i s c e r t a i n t h a t t h e r e w i l l be b o t h m a j o r and minor t e c h n o l o g i c a l b r e a k t h r o u g h s or d i s c o v e r i e s t h a t w i l l have a great t o t a l impact on our a g r i c u l t u r e and c r o p p r o d u c t i o n of the f u t u r e . We are j u s t now s e e i n g some of the e a r l y p r o d u c t s of the p i o n e e r i n g e f f o r t s i n t r o d u c e d f o r f i e l d t r i a l s or i n i t i a l a p p l i c a t i o n s . Major b r e a k t h r o u g h s are b e i n g made which w i l l make i t t e c h n o l o g i c a l l y p o s s i b l e t o produce g e n e t i c a l l y e n g i n e e r e d organisms e a r l i e r than many had p r e d i c t e d . However, the f i n a l and u l t i m a t e c r o p s or organisms w i l l l i k e l y r e q u i r e a l o n g e r time t o d e v e l o p or p e r f e c t than many b i o t e c h n o l o g i s t s expect. Status

of the

Technology

One o f the reasons f o r the g r e a t amount of time needed b e f o r e major p r o d u c t s of b i o t e c h n o l o g y w i l l be ready f o r commercial use i s the p r e s e n t l i m i t a t i o n s o f b i o t e c h n o l o g y i t s e l f . For example, w h i l e s c i e n t i s t s have a c h o i c e of ways t o e n g i n e e r new p l a n t s , a t h r e e - s t a g e p r o c e s s i s r e q u i r e d , not a l l s t e p s of which can r e a d i l y be done w i t h major c r o p s , such as c o r n and soybeans. The f i r s t s t a g e i n e n g i n e e r i n g a new c r o p p l a n t i s t o e s t a b l i s h a c e l l c u l t u r e of the p l a n t so t h a t f o r e i g n DNA can be inserted. T i s s u e from any p l a n t or seed p a r t can be used t o e s t a b l i s h the i n i t i a l c e l l c u l t u r e . T h i s donor t i s s u e i s put onto a s y n t h e t i c growth medium, where i t s t a r t s d i v i d i n g and forms a type of wound t i s s u e c a l l e d c a l l u s . This c a l l u s i s e x c i s e d from the donor t i s s u e and m a i n t a i n e d i n c u l t u r e i n d e f i n i t e l y by r e p l e n i s h i n g i t s medium. By m a n i p u l a t i n g the medium, p a r t i c u l a r l y the hormone c o n t e n t , s c i e n t i s t s can induce c a l l u s t i s s u e t o r e g e n e r a t e i n d i v i d u a l organs or a complete plant. In the second s t a g e , the d e s i r e d g e n e t i c m a t e r i a l or f o r e i g n DNA must be t r a n s f e r r e d i n t o the c e l l i n - c u l t u r e , and the c e l l s t h a t accept the new DNA must be s e p a r a t e d from the r e m a i n i n g unchanged c e l l s . There are s e v e r a l ways t o i n s e r t new g e n e t i c m a t e r i a l i n t o p l a n t c e l l s once they have been e s t a b l i s h e d i n the c u l t u r e medium. The f i r s t and major method o f DNA t r a n s f e r i s c e l l f u s i o n . T h i s can be a c c o m p l i s h e d s e v e r a l ways, a l l of which r e q u i r e u s i n g p l a n t p r o t o p l a s t s , or p l a n t c e l l s whose c e l l w a l l s have been d i g e s t e d away by enzymes. A f t e r the p l a n t c e l l w a l l s have been s t r i p p e d o f f , the c o n t e n t s o f a d j a c e n t c e l l s can be f o r c e d t o combine or fuse i n a high-pH, h i g h - c a l c i u m medium. Other methods of f o r c i n g c e l l f u s i o n i n c l u d e p o l y e t h y l e n e g l y c o l t r e a t m e n t o f p l a n t membranes and a p r o c e s s c a l l e d e l e c t r o p o r a t i o n , i n which p u l s e s o f e l e c t r i c i t y cause s m a l l pores



1.

LEBARON

Status and Overview

to be c r e a t e d i n the p l a n t membranes, t h r o u g h which l a r g e m o l e c u l e s ( e . g . , DNA) e n t e r . While h e a l i n g , the p r o t o p l a s t membranes fuse t o g e t h e r . T h i s p r o c e s s , which has been used f o r a number o f years on animal c e l l c u l t u r e s , s h o u l d work w i t h p l a n t cells. Another s i m i l a r method t h a t has had l i m i t e d t e s t i n g employs d i r e c t m i c r o i n j e c t i o n o f DNA i n t o the n u c l e u s o f p r o t o p l a s t s w i t h p a r t l y r e g e n e r a t e d c e l l w a l l s . A f o u r t h method f o r DNA t r a n s f e r i n v o l v e s u s i n g s y n t h e t i c liposome c a r r i e r s t o move t h e DNA. Liposomes are l a b o r a t o r y - m a d e membrane packages i n t o which DNA can be i n c o r p o r a t e d . A liposome can be fused w i t h a p l a n t p r o t o p l a s t by the same t e c h n i q u e s used f o r c e l l f u s i o n . However, none o f the methods o f t r a n s f o r m i n g p r o t o p l a s t s can y e t be u s e f u l f o r corn or c e r e a l c r o p s as no good system e x i s t s f o r r e g e n e r a t i n g p l a n t s from monocot p r o t o p l a s t s . A f i f t h and a n o t h e r f r e q u e n t l y used as w e l l as s u c c e s s f u l method r e q u i r e s u s i n g a v e c t o r such as a b a c t e r i u m t o c a r r y the f o r e i g n DNA i n t o the c e l l . The most prominent v e c t o r used i n g e n e t i c e n g i n e e r i n g experiments has been the b a c t e r i u m A g r o b a c t e r i u m t u m e f a s c i e n s , w h i c h causes crown g a l l d i s e a s e i n many d i c o t p l a n t s . This b a c t e r i u m a l r e a d y has a mechanism f o r i n v a d i n g a p l a n t c e l l and i n t r o d u c i n g i t s DNA i n t o a p l a n t n u c l e u s . Genetic engineers i n s e r t the d e s i r e d g e n e t i c i n f o r m a t i o n i n t o t h e A g r o b a c t e r i u m DNA so t h a t when the b a c t e r i u m i n f e c t s t h e c e l l , i t c a r r i e s along the new DNA. U n f o r t u n a t e l y , t h i s b a c t e r i u m does not invade monocot c r o p s , such as corn and c e r e a l s , so we must s e a r c h f o r o t h e r s u i t a b l e v e c t o r s t h a t w i l l i n f e c t monocot p l a n t s and c a r r y the f o r e i g n DNA i n t o them. In the t h i r d s t a g e , the t r a n s f o r m e d c e l l s must be regene r a t e d i n t o whole p l a n t s , which must then be t e s t e d t o ensure t h a t the e n g i n e e r e d t r a i t has been s u c c e s s f u l l y i n h e r i t e d and w i l l be passed on t o the next g e n e r a t i o n v i a the seed. There a r e two methods f o r g e t t i n g c e l l s t h a t have a c c e p t e d f o r e i g n DNA t o regenerate. In one, c a l l e d o r g a n o g e n e s i s , the c u l t u r e d t i s s u e produces a shoot apex, which i s a l l o w e d t o grow. A f t e r a c e r t a i n p o i n t , the shoot i s cut from the c a l l u s t i s s u e and put i n t o a d i f f e r e n t c u l t u r e medium t h a t a l l o w s r o o t i n g . When the shoot has r o o t e d , i t i s put i n t o s o i l and grows i n t o a mature plant. The o t h e r method o f r e g e n e r a t i o n i s s o m a t i c embryo format i o n , which can take advantage o f s o m a c l o n a l v a r i a t i o n s , o r the inherent genetic d i v e r s i t y a v a i l a b l e i n plant c e l l s . Somaclonal v a r i a t i o n r e f e r s t o the o b s e r v a t i o n t h a t i n d i v i d u a l c u l t u r e d p l a n t c e l l s do not always d i s p l a y the same g e n e t i c i n f o r m a t i o n ( e . g . , h e r b i c i d e t o l e r a n c e ) as the p l a n t from which the c e l l s were d e r i v e d . In a d d i t i o n , the a b i l i t y t o e x p l o i t somaclonal v a r i a t i o n does not r e q u i r e the i d e n t i f i c a t i o n and t r a n s f e r o f i n d i v i d u a l genes. Somatic embryos a r e formed from nonreproductive c e l l s . By m a n i p u l a t i n g the medium, r e s e a r c h e r s can o f t e n get the somatic embryos t o grow i n t o complete p l a n t s . However, these r e g e n e r a t e d p l a n t s have o f t e n been s e n s i t i v e t o h u m i d i t y , temperature changes, and f u n g a l i n f e c t i o n s . When the somatic embryo has grown t o m a t u r i t y , seed i s c o l l e c t e d and checked t o see i f the d e s i r e d t r a i t has been i n h e r i t e d .


11


12


Even a f t e r the g e n e t i c e n g i n e e r has s u c c e s s f u l l y completed h i s work and has c o n f i r m e d t r a n s f e r of t h e g e n e t i c t r a i t i n t o a new c r o p , much more work i s s t i l l ahead. Some o f t h i s , such as c o n f i r m i n g t h a t the g e n e t i c t r a n s f e r i s s t a b l e o r c o m p a t i b l e o v e r many g e n e r a t i o n s , t h a t t h e t r a i t i s e x p r e s s e d o n l y i n the proper organ, at the proper t i m e , and has no d e l e t e r i o u s e f f e c t s on the r e s t o f the p l a n t ' s genome e x p r e s s i o n or p h y s i o l o g y , can be c a r r i e d out by, o r i n c o o p e r a t i o n w i t h , the g e n e t i c e n g i n e e r . However, much time, r e s e a r c h , and t e s t i n g w i l l be r e q u i r e d by the b r e e d e r , p h y s i o l o g i s t , agronomist and o t h e r s a f t e r the b i o t e c h n o l o g i s t s or g e n e t i c e n g i n e e r s complete t h e i r j o b . A l t h o u g h the c o o p e r a t i o n and i n t e r a c t i o n between b i o t e c h n o l o g y and these o t h e r d i s c i p l i n e s have g r e a t l y improved i n recent y e a r s , t h e r e i s s t i l l a need f o r i n c r e a s e d t r u s t , c o l l a b o r a t i o n , and j o i n t r e s e a r c h at many i n s t i t u t i o n s . As s t a t e d by Hugh B o l l i n g e r o f N a t i v e P l a n t s , "Commercial p r o s p e c t s f o r new p l a n t p r o d u c t s may be i n h i b i t e d by the d i f f i c u l t i e s o f s c a l i n g up p r o d u c t i o n and by the number of t a l e n t e d people t r a i n e d i n p l a n t g e n e t i c s " ( 1 3 ) . J a w o r s k i f u r t h e r emphasized t h a t , "The most fundamental problem i n a p p l y i n g g e n e t i c e n g i n e e r i n g t o a g r i c u l t u r e i s a l a c k of b a s i c b i o c h e m i c a l knowledge about p l a n t s . We need t o spend a l o t more t i m e — and t h i s i s where I t h i n k we w i l l see a g r e a t d e a l o f a c t i v i t y i n the next f i v e t o t e n y e a r s — on i d e n t i f y i n g a g r o n o m i c a l l y important t r a i t s and the genes t h a t r e g u l a t e those t r a i t s . I f we cannot do t h i s , we are not g o i n g t o be v e r y s u c c e s s f u l i n r e a l l y making the agronomic improvements t h a t we d e s i r e t o make. We j u s t don't have enough knowledge yet t o u n d e r s t a n d how t o r e g u l a t e at w i l l , and i n a c o n t r o l l e d f a s h i o n , the e x p r e s s i o n o f a gene. There i s a l o t of b a s i c r e s e a r c h t h a t has t o be done i n p a r a l l e l w i t h the a p p l i e d r e s e a r c h i f we are g o i n g t o be s u c c e s s f u l i n moving the t e c h n o l o g y from the l a b o r a t o r y i n t o the f i e l d " (2^). The r e s e a r c h needed t o a c q u i r e t h i s knowledge r e q u i r e s b o t h g r e a t e r c o o p e r a t i o n between p l a n t m o l e c u l a r b i o l o g i s t s and t r a d i t i o n a l p l a n t b r e e d e r s and a commitment by the f e d e r a l government t o fund t h i s k i n d of i n t e r d i s c i p l i n a r y e f f o r t . Biotechnology

Patents

The problems o f patent p r o t e c t i o n , i n a d d i t i o n t o governmental r e g u l a t i o n s , add elements of r i s k to a l l g e n e t i c e n g i n e e r i n g projects. The commercial p o t e n t i a l f o r p r o d u c t s of b i o t e c h n o l o g y was made p o s s i b l e or at l e a s t g r e a t l y enhanced by two landmark d e c i s i o n s i n v o l v i n g p a t e n t s i n r e c e n t y e a r s . The f i r s t was i n 1980 when the Supreme Court d e c i d e d t h a t u n i c e l l u l a r organisms c o u l d be p a t e n t e d (Diamond v s . C h a k r a b a r t y #447, U.S. 303). This new r u l i n g went on t o i n c l u d e p l a n t s , seeds and t i s s u e c u l t u r e s too, i n d i c a t i n g t h a t m u l t i - c e l l e d , complex h i g h e r l i f e forms w i t h stems, f r u i t s , f l o w e r s , e t c . , c o u l d a l s o be p a t e n t e d . The second landmark d e c i s i o n was i n O c t o b e r , 1985, v e r y soon a f t e r the symposium, when the U.S. Patent A p p e a l s Board awarded M o l e c u l a r G e n e t i c s a patent f o r a g e n e t i c a l l y m o d i f i e d p l a n t . T h i s g r a n t e d f u l l patent p r o t e c t i o n t o a corn p l a n t , seeds and t i s s u e c u l t u r e



LEBARON

Status and Overview

t h a t M o l e c u l a r G e n e t i c s s c i e n t i s t s m o d i f i e d t o overproduce the amino a c i d t r y p t o p h a n , a c h a r a c t e r i s t i c t h a t w i l l make c o r n more n u t r i t i o u s f o r animals and may e l i m i n a t e t h e need f o r d i e t a r y supplements. I t c o u l d be t h e green l i g h t t h e b i o t e c h n o l o g y i n d u s t r y has been l o o k i n g f o r t o c r e a t e new s t r a i n s o f p l a n t s , and c e r t a i n l y p l a c e s p a t e n t a b i l i t y o f e n g i n e e r e d p l a n t s i n a new light. P r i o r t o t h i s a c t i o n , the o n l y way t o o b t a i n p a t e n t s on p l a n t s propagated by seed was t o do so on i n d i v i d u a l v a r i e t i e s through USDA. The Patent O f f i c e had p r e v i o u s l y i n s i s t e d t h a t i t couldn't protect c h a r a c t e r i s t i c s of p l a n t s . I f Molecular G e n e t i c s had gone t o t h e USDA t o patent i t s c h a r a c t e r i s t i c , i t would have had t o f i l e s e p a r a t e a p p l i c a t i o n s f o r each d i f f e r e n t k i n d o f corn v a r i e t y . Now covered by g e n e r a l patent law, g e n e t i c engineering for overproduction of tryptophan i s protected i n a l l c o r n p l a n t s . The s t a n d a r d f o r p a t e n t a b i l i t y o f b i o t e c h n o l o g y p r o d u c t s now seems t o be t h a t i t o n l y has t o be n o v e l , u s e f u l , and not obvious ( 1 4 ) . On t h e o t h e r hand, the p r o c e s s o f a p p l y i n g f o r such p r o t e c t i o n i s a t h o r n y i s s u e f o r a g r i b u s i n e s s . Some o f t h e d i f f i c u l t i e s i n c l u d e l a c k o f a v a i l a b l e and knowledgeable manpower, l a c k o f t r a i n i n g w i t h i n t h e p a t e n t a g e n c i e s , and how t o b e s t prepare p a t e n t s f o r optimum coverage. Most o f the c u r r e n t examiners o f new b i o t e c h n o l o g i c a l a p p l i c a t i o n s have a background i n c h e m i s t r y . F r e q u e n t l y , they e v a l u a t e n o v e l t i e s i n a g r i b i o t e c h i n a manner d i f f e r e n t from t h a t used by b i o l o g i s t s . Besides, t h e y can o n l y spend about seven t o e i g h t hours w i t h an a p p l i c a t i o n b e f o r e they have t o d r a f t a judgment. C l e a r l y p r e s e n t e d a p p l i c a t i o n s which c l a i m a narrow p r o t e c t i o n have t h e b e s t chance o f b e i n g approved. Broad c l a i m s may be r e j e c t e d and c a r r y t h e danger o f b e i n g t i e d up i n i n t e r f e r e n c e s u i t s . These are c o s t l y and may l a s t two t o t h r e e y e a r s . N o n e t h e l e s s , b i o t e c h n o l o g y i s here t o s t a y and i s a welcome b l e s s i n g i n p r o v i d i n g new, b e t t e r and f a s t e r means t o develop c r o p s , animals and food f o r our f u t u r e and growing p o p u l a t i o n s . P r a c t i c a l C o n s i d e r a t i o n s i n t h e M a r k e t i n g and A p p l i c a t i o n s of B i o t e c h n o l o g y P r o d u c t s The f i n a l f a c t o r s n e c e s s a r y f o r a product o f b i o t e c h n o l o g y or any s c i e n c e t o be s u c c e s s f u l a r e economics, p r o f i t a b i l i t y and farmer a c c e p t a n c e . The u s e r may not understand o r c a r e about the degree o f s c i e n t i f i c s o p h i s t i c a t i o n o r what marvelous d i s c o v e r i e s have l e d t o t h e f i n a l p r o d u c t . He w i l l o n l y ask: " I s i t s i m p l e t o use, e c o n o m i c a l and w i l l i t make me a p r o f i t ? " We must a l s o r e a l i z e t h a t i n most c a s e s , t h e p r o d u c t s o f b i o t e c h n o l o g y w i l l e i t h e r be i n t r o duced i n c o m p e t i t i o n w i t h c h e m i c a l s o r o t h e r p r o d u c t s t h a t a r e a l r e a d y accepted and a r e s a t i s f y i n g most needs, o r they w i l l e n t e r new areas and markets where n o t h i n g i s p r e s e n t l y a v a i l a b l e or s a t i s f a c t o r y . Both c o n d i t i o n s p r e s e n t s e r i o u s economic r i s k s and a c c e p t a n c e problems.



14

BIOTECHNOLOGY IN AGRICULTURAL CHEMISTRY With few e x c e p t i o n s , pest c o n t r o l i n modern-day a g r i c u l t u r e i s based on c h e m i c a l s . To s u c c e s s f u l l y i n t r o d u c e a b i o l o g i c a l l y - b a s e d pest management system i n t o a g r i c u l t u r e r e q u i r e s t h a t a minimum o f new t e c h n o l o g y or requirements be p l a c e d on the d i s t r i b u t i o n system. T h i s a p p l i e s t o a l l parameters o f the p r o d u c t : s h e l f l i f e , h a n d l i n g , c o m p a t i b i l i t y w i t h o t h e r p e s t i c i d e s , e f f i c a c y , and above a l l , c o s t per a c r e t r e a t e d . A b i o l o g i c a l l y based product must be a b l e t o f i l l a r e a l need i n the m a r k e t p l a c e and must be a b l e t o do i t through the e s t a b l i s h e d channels now used by c h e m i c a l s . U l t i m a t e success w i l l s t i l l be determined by how the p l a n t or organism a c t u a l l y performs i n the f i e l d . G e n e t i c a l l y e n g i n e e r e d c r o p s can o n l y succeed i f they improve the farmer's b a l a n c e s h e e t . G e n e t i c e n g i n e e r i n g cannot a l t e r b a s i c r u l e s o f p h y s i o l o g y . I f improving the n u t r i t i o n a l q u a l i t y of a c e r e a l r e s u l t s i n s u b s t a n t i a l y i e l d l o s s e s because o f u n d e r l y i n g b a s i c p r o c e s s e s , g e n e t i c e n g i n e e r i n g w i l l not circumvent the problem. Genetic engineering p o t e n t i a l l y o f f e r s s i g n i f i c a n t o p p o r t u n i t i e s i n crop a g r i c u l t u r e , but o n l y i f b a s i c economics are not a d v e r s e l y a f f e c t e d by the changes i n d u c e d i n the p l a n t . Even t h e n , a s u c c e s s f u l g e n e t i c e n g i n e e r i n g program may s t i l l r e q u i r e b a s i c c l a s s i c a l b r e e d i n g t e c h n o l o g y t o i n t r o d u c e the new t r a i t i n t o numerous v a r i e t i e s . F u r t h e r , i n t r o d u c i n g a new v a r i e t y i n t o the m a r k e t p l a c e r e q u i r e s t h a t i t s c h a r a c t e r i s t i c s be determined v i a f i e l d t e s t i n g . T h e r e f o r e , even i f a s u b s t a n t i a l l y improved g e n e t i c a l l y engineered v a r i e t y was a v a i l a b l e at the greenhouse l e v e l now, years o f f i e l d t e s t i n g would be r e q u i r e d b e f o r e widespread commercial u t i l i z a t i o n can o c c u r ( 1 5 ) .

Literature Cited 1) 2) 3) 4) 5) 6) 7) 8)

9) 10)

11)

Hardy, R. W. F . ; Glass, D. J . Issues on Science and Technology 1985, 6, 69-82. Olson, S. "Biotechnology - An Industry Comes of Age"; National Academy Press: Washington, D.C. 1986; p. 120. Valiulis, D. Agrichemical Age 1984, 28(8), 53-60. Kaufman, R. Personal Communication, 1986. Rifkin, J . "Declaration of a Heretic"; Rout ledge and Kegan Paul: Boston, MA, 1985; p. 140. Lappe, M. "Broken Code: The Exploitation of DNA"; Sierra Club Books: San Francisco, CA, 1985; p. 354. B r i l l , W. Science 1985, 227, 381-4. LeBaron, H. M.; Gressel, J . In "Herbicide Resistance in Plants"; LeBaron, H. M.; Gressel, J., Eds.; John Wiley and Sons: New York, 1982; pp. 349-362. McWhorter, C. G. Weed Science 1984, 32, 850-5. Shaw, W. C. In "Agricultural Chemicals of the Future"; The ARS National Research Program. Hilton, J . L. Ed.; BARC Symposium No. 8, Rowman and Allanheld: Totowa, NJ, 1984. Dempster, C. Industrial Chemical News 1986, 7(7),20-1.


1.

LEBARON

12) 13) 14) 15) 16) 17) 18) 19)


20)

Status and Overview

Arntzen, C. J.; Pfister, K.; Steinback, Κ. E. In "Herbicide Resistance in Plants"; LeBaron, H. M.; Gressel, J., Eds.; John Wiley and Sons: New York, 1982; pp. 185-214. Lewis, R. High Technology 1986, 6(5), 46-53. Weber, J . Genetic Engineering News 1986, 6(7), 3. Kempner, D. H.; Schnell, G. W.; Wolnak, B. Genetic Engineering News 1986, 6(1), 18-19. Miller, C. Genetic Engineering News 1986, 6(7), 41. Cannon, D. R. Industrial Chemical News 1986, 7(8), 8. McCormick, D. Bio/Technology 1985, 3(5), 429-35. Kearney, P. C.; Karns, J . S.; Mulbry, W.; Tomasek, P.; Shelton, D. J . Agr. Food Chem. 1986, In press. Robotti, Κ. M.; Sharp, J . K.; Ehrmann, P. R.; Brown, L. J.; Hermann, B. W. J . Agr. Food Chem. 1986, In press.

RECEIVED November 26,

1986


15

Status and Overview of Biotechnology in ... - ACS Publications

Recommend Documents