The document summarizes the history and process of anther and pollen culture. Key developments include the first observation of pollen grain proliferation in 1953, and the first reports of embryo development from microspores in 1964 and 1967. Pollen or microspore culture involves culturing pollen grains, preferably at the uninucleated stage, on nutrient medium to develop into haploid embryoids or callus tissue that give rise to haploid plantlets. The production of haploid plants exploits the totipotency of microspores and diverts their development away from becoming male gametes.

2. HISTORY
• W.TULECKE (1953)
FIRST OBSERVED THAT MATURE POLLEN GRAINS OF GINKGO BILIBA CAN BE INDUCED TO PROLIFERATE IN
CULTURE TO FORM HAPLOID CALLUSS
• S.GUHA AND S.C.MAHESWARI (1964)
FIRST REPORTED THE DIRECT DEVELOPMENT OF EMBRYOS FROM MICROSCOPE OF DATURA INNOXIA BY THE
CULTURE OF EXCISED ANTHER
• J.P.BOURGIN AND J.P.NITSCH (1967)
• OBTAINED COMPLETE HAPLOID PLANTLET FROM ANTHER CULTURE OF NICOTIANA TABACUM .

3. POLLEN CULTURE
• POLLEN OR MICROSPORE CULTURE IS AN IN VITRO
TECHNIQUE BY WHICH THE POLLEN GRAINS PREFERABLY
AT THE UNINUCLEATED STAGE, ARE SQUEEZED OUT
ASEPTICALLY FROM THE INTACT ANTHER AND THEN
CULTURED ON NUTRIENT MEDIUM.
• THE MICROSPORES DEVELOP INTO HAPLOID EMBRYOIDS
OR CALLUS TISSUE THAT GIVE RISE TO HAPLOID
PLANTLETS BY EMBRYOGENESIS OR ORGANOGENESIS.

4. ANDROGENESIS
ANDROGENESIS IS THE IN VITRO DEVELOPMENT OF
HAPLOID PLANTS ORIGINATING FROM TOTIPOTENT
POLLEN GRAINS THROUGH A SERIES OF CELL DIVISION
AND DIFFERENTIATION.
IT IS OF TWO TYPES:
1) DIRECT ANDROGENESIS
2) INDIRECT ANDROGENESIS

5. 1)DIRECT ANDROGENEIS:-
THE MICROSPORES BEHAVES LIKE A ZYGOTE AND UNDERGOES CHANGE TO FORM ENBRYOID WHICH
ULTIMATELY GIVE RISE TO A PLANTLET.
2) INDIRECT ANDROGENESIS:
THE MICROSPORES DIVIDE REPEATEDLY TO FORM A CALLUS TISSUE WHICH DIFFERENTIATES INTO
HAPLOID PLANTLETS.

6. PRINCIPLES OF POLLEN CULTURE
• THE PRODUCTION OF HAPLOID PLANTS IS TO
EXPLOIT THE TOTIPOTENCY OF MICROSPORE.
• IN THIS PROCESS THE NORMAL DEVELOPMENT AND
FUNCTION OF THE POLLEN CELL TO BECOME A MALE
GAMETE IS STOPPED AND IS DIVERTED FORCELY TO A
NEW METABOLIC PATHWAY FOR VEGETATIVE CELL
DIVISION.

7. METHOD OF POLLEN CULTURE
• ANTHER COLLECTED FROM FLOWER BUDS AND POLLEN GRAINS ARE ISOLATED AND ABOUT 50 ANTHERS ARE PLACED IN
SMALL STERILE BEAKER CONTAINING 20 ML OF LIQUID BASAL MEDIUM (MS OR WHITE OR NITSCH)
• ANTHERS ARE THEN PRESSED AGAINST THE SIDE OF BEAKER WITH THE STERILE GLASS ROD TO SQUEEZE OUT THE
POLLENS.
• THE HOMOGENIZED ANTHERS ARE THEN FILTERED THROUGH A NYLON SIEVE TO REMOVE THAT THE ANTHER TISSUE
DEBRIS.
• THE FILTRATE OR POLLEN SUSPENSION IS THEN CENTRIFUGED AT LOW SPEED (500-800 RPM/MIN) FOR FIVE MINUTES.
THE SUPERNATANT CONTAINING FINE DEBRIS IS DISCARDED AND PELLET OF POLLEN IS SUSPENDED IN FRESH LIQUID
MEDIUM AND WASHED TWICE BY REPEATED CENTRIFUGATION AND RESUSPENDED IN FRESH LIQUID MEDIUM.

8. • A 2.5 ML OF POLLEN SUSPENSION(USUALLY 10’ TO10 4/ML)
IS PIPETTED OFF AND IS SPREAD IN 5 CM PETRIDISH.
POLLENS ARE BEST GROWN IN LIQUID MEDIUM BUT, IF
NECESSARY, THEY CAN BE GROWN BY PLATING VERY SOFT
AGAR ADDED MEDIUM.
• PETRIDISHES ARE INCUBATED AT 27-30°C UNDER LOW
INTENSITY OF WHITE COOL LIGHT (500 LUX, 16 HR).
• YOUNG EMBRYOIDS CAN BE OBSERVED AFTER 30 DAYS. THE
EMBRYOLDS ULTIMATELY GIVE RISE TO HAPLOID PLANTLETS.

9. ADVANTAGES OF POLLEN CULTURE
• UTILITY OF ANTHER AND POLLEN CULTURE FOR BASIC RESEARCH CYTOGENETIC STUDIES.
• STUDY OF GENETIC RECOMBINATION IN HIGHER PLANTS.
• STUDY OF MODE OF DIFFERENTIATION FROM SINGLE CELL TO WHOLE ORGANISM.
• STUDY OF FACTORS CONTROLLING THE POLLEN EMBRYOGENESIS OF HIGHERPLANTS.
• FORMATION OF DOUBLE HAPLOID THAT ARE HOMOZYGOUS AND FERTILE.

10. ADVANTAGES OF POLLEN CULTURE
• FOR MUTATION STUDY.
• FOR PLANT BREEDING AND CROP IMPROVEMENT. TO OBTAIN THE SECONDARYMETABOLITES.
• EXAMPLE:‘HYOSCYAMUS NIGRER’ OBTAIN BY ANTHER
• CULTURE HAVING HIGHER ALKALOID CONTENT.
• HAPLOIDS ARE USED IN MOLECULAR BIOLOGY AND GENETIC ENGINEERING.