INDIA
ITG/13
Date : 8-7-2003
National Guidelines
for the Conduct of Test for
Distinctness, Uniformity and Stability
Rape seed and Mustard
(Brassica)

Contents
I.            Subject
II.           Material required
III.          Conduct of tests  
IV.         Methods and observations  
V.          Grouping of varieties  
VI.         Characteristics and symbols  
VII.        Table of characteristics  
VIII.       Explanations on the table of characteristics  
IX.         Technical questionnaire  

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  I.   Subject

These test guidelines apply to all varieties, hybrids and parental lines of Rape seed and Mustard, comprising yellow sarson (Brassica rapa var. yellow sarson), brown sarson (B. rapa var. brown sarson), toria (B.rapa var. toria), Indian mustard (B. juncea L.), karan rai (B. carinata), gobhi sarson (B. napus L.) and black mustard (B. nigra L.) 

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II.   Material required

  1. The Plant Variety Protection (PVP) Authority decides when, where and in what quantity and quality the seed material required for testing the variety is to be delivered.  Applicants submitting material from a country other than India must make sure that all customs formalities are complied with.  The minimum quantity of seed to be supplied by the applicant should be :  

    Variety and hybrid     -           500 gram (in one submission only)

    Parental lines           -         100  gram  

  2. The seed should meet the minimum requirements for germination capacity, moisture content and physical purity prescribed for certified seed in India . Especially for storage, which requires a higher standard, the applicant should state, the actual germination capacity which should be as high as possible.
     

  3. The seed material must not have undergone any treatment unless the competent authorities allow or request such treatment.  If it has been treated, full details of the treatment must be given.  

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III.   Conduct of tests

  1. The minimum duration of tests should normally be two independent similar growing seasons with reference to the ecosystem of the variety submitted for DUS test.
     

  2. The test should normally be conducted at two test locations. If any important characteristics of the variety can not be seen at these places, the variety may be tested at an additional place.  
      

  3. The field tests should be carried out under conditions ensuring normal growth.  The size of the plots should be such that plants or parts of plants may be removed for measurement and counting without prejudice to the observations which must be made upto the end of the growing period. Each test should include about 400 plants, which should be divided among three replications. Separate plots for observation and for measurement can only be used if they have been subjected to similar environmental conditions.  
      

  4. Test plot design  

    Number of rows : 5
    Row length : 5 m
    Row to row distance : 45 cm
    Plant to plant distance : 15 cm
    Number of replications : 3


  5. Observations should not be recorded on plants in border rows.  
     

  6.  Additional tests for special purpose may be established.  

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IV.   Methods and observations

  1. The characteristics described in the table of characteristics should be used for the testing of varieties, inbred lines and hybrids for DUS.  
      

  2. For the assessment of distinctness and stability observations should be made on 60 plants or parts of 60 plants which should be divided among 3 replications (20 plants per replication).  
      

  3. For the assessment of uniformity of characteristics on the plot as a whole (visual assessment by a single observation of a group of plants or parts of plants), of parental lines a population standard of 1% with an acceptance probability of 95% and for varieties and hybrids, a population standard of 5% with an acceptance probability of at least 95% should be applied.
     

  4. To assess distinctness of hybrids, a prescreening system on the basis of the parental lines and the formula may be established according to the following recommendations.  

    a)     description of parental lines according to the Test Guidelines

    b)     check of the originality of those parental lines in comparison with the reference collection, based on the characteristics in Chapter VII in order to screen the closest inbred lines

    c)      check of the originality of the hybrids formula in comparison with those of hybrids in common knowledge, taking into account the closest inbred lines.

    d)     Assessment of the distinctness at the hybrid level of varieties with a similar formula.  
      

  5. For the assessment of colour characteristics, it is recommended that Royal Horticultural Society (RHS) colour chart should be used.  
      

  6. Unless otherwise indicated, all observations on the foliage should be made on  fully developed leaves in the rosette.  
     

  7. Unless otherwise indicated, all observations on siliquas should be made on  fully developed  siliqua from the lower third on the main stem.  

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V.   Grouping of varieties

  1. The collection to be grown should be divided into groups to facilitate the assessment of distinctness. Characteristics that are suitable for grouping purposes are those which are known from experience not to vary, or to vary only slightly, within a variety and which in their various states are fairly evenly distributed within the collection.  
      

  2. It is recommended that the competent authorities use the following characteristics for grouping varieties.

                                                              i.            Leaf : lobes (Characteristic 6)

                                                            ii.            Time of flowering (Characteristic 12)

                                                          iii.            Anther : production of pollen (Characteristic 16)

                                                           iv.            Plant : main shoot length (Characteristic 17)

 

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VI. Characteristics and symbols

  1. To assess distinctness, uniformity and stability, the characteristics and their states as given in the Table of Characteristics should be used. 
      

  2. Notes (1 to 9), for the purposes of electronic data processing, are given opposite the states of the different characteristics.  
     

  3. Legend :

    (*)        Characteristics that should be used every growing period for the examinations of all varieties and should always be included in the description of the variety, except when the state of expression of a preceding characteristic or regional environmental conditions render this impossible.

    (+)       See Explanations on the table of characteristics in Chapter VIII.  

  4. The optimum stage of development of plant growth for the assessment of each characteristics is indicated by a number in the sixth column of table of characteristics. The stages of development denoted by each number are described below : 
     
    According to Berkenkamp, 1973  

    KEY              GENERAL DESCRIPTION  
    0 Germination
    00 Dry Seed
    10 Seedling Growth
    11 Appearance of cotyledons
    13 Cotyledons of expended
    15 1 Leaf stage
    17 2 Leaf stage
    19 3 Leaf stage
    20 Rosette
    21 4 Leaf stage
    22 5 Leaf stage
    23 6 Leaf stage
    24 7 Leaf stage
    25 8 Leaf stage
    26  9 - 11 Leaf stage
    27 12 or more leaves are completely developed
    30 Stem elongation
    31 Distance between cotyledons and vegetation point more than 5 cm
    35 Distance between cotyledons and vegetation point is more than 15 cm
    39 Distance between cotyledons and vegetation point is more than 25 cm
    50 Bud formation
    51 Terminal bud is present, not raised above leaves
    53 Terminal bud is raised above level of leaves
    57 Pedicel are elongating
    59 Buds are yellowing
    60 Flower
    61 First open bud terminal raceme
    62 Few buds are open on terminal raceme
    64 Full flower, lower siliqua are elongating
    65 Lower siliques are starting to fill, less than 5% of buds are not yet open
    67 Seed in lower siliquaes are enlarging, all buds are open
    70 Siliqua
    71 Seed in lower siliquaes are in full size translucent
    75 Seed in lower siliquaes are green opaque
    79 All seed of  siliquaes on terminal raceme are dark
    80 Maturation
    81 seed in lower Siliquaes on terminal raceme show brown areas
    85 Seed in upper Siliquaes show brown areas
    89 Brown Siliquaes are brittle, stems are dry

     

  5. Type of assessment of characteristics indicated in column 7 of table of characteristics is as follows :  

    MG : Measurement by a single observation of a group of plants or parts of plants

    MS : Measurement of a number of individual plants or parts of plants

    VG : Visual assessment by a single observation of a group of plants or parts of plants

    VS : Visual assessment by observations of individual plants or parts of plants

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VII. Table of characteristics

S.No. Characteristics States Notes Example Varieties Stage of observation Type of assessment

1.
(+)

Seed: erucic Acid

low (<2 %)
medium (2-25 %)
high (26 -50 %)


very high (>50 %)

3
5
7


9

 

Bhawani, GSL1, Kiran, Pusa Bold,RH30
PT303,GSL2, Jhumka

00

MG

2.
(+)

Seed : glucosinolate

low (<31 μm/g)
medium (31-60 μm/g)

high (61-90 μm/g )


very high (>90μm/g )

3
5

7


9

 
TH68,Sheetal, RCC4
PT303,GSL1, Pusa Gold, RTM314, Varuna
Bhawani, GSL2, Jhumka, Kiran,PCR7

00

MG

3.
(*)
(+)

Seed : oil content

low (< 38 %)

medium (38-42 %)


high (43-46 %)

very high (>46)

3

5


7

9

PC5,RTM314, Kranti
Bhawani,GSL1, Pusa Gold, Kiran, Varuna
PT303, Jhumka, RN393

00

MG

4.

Leaf : hairiness

absent


sparse
dense

1


3
7

Bhawani,GSL1, Pusa Gold, Kiran, Varuna, RTM 314

6-12

VS

5.
(*)

Leaf : colour

light green
medium green  

dark green
purple green  
purple  

1
2

3
4
5

 
Bhawani, Pusa Gold, RTM314  
GSL1, Kiran

23-27

VG

6.
(+)

Leaf : lobes

absent  
present

 

1
9

Jhumka  
Bhawani, PT303, GSL1,Pusa Gold, Kiran, Varuna

23-27

VS

7.
(*)
(+)

Leaf : number of lobes

few (1-2)  
medium (3-5)  
many (>5)

 

3
5
7

 

Bhawani, PT303, GSL1,Pusa Gold, Kiran, Varuna  

23-27

MS

 

8.
(*)
(+)

Leaf : dentation of margin

entire  



auriculate  

lyrate  
pointed

1



3

5
7

PT303, GSL1, Pusa Gold, Kiran, RTM 314, 
GSL1.
Jhumka,KBS3, Rohini, GM1, CS52  

Pusa Bold, Varuna

23-27

VS

 

9.
(+)

Leaf : length

short (<15 cm)  

medium (15-20 cm)
long (>20 cm)

3

5
7

Bhawani, PT507, PT303, TH68, TL15
Pusa Gold
GSL1, Kiran, RTM 314, Varuna

23-27

MS

 

10.
 (+)

Leaf : width

narrow (<7 cm)  


medium (7-8 cm)  
broad (>8 cm)

3


5
7

Bhawani, PT507, PT303, TH68, Jhumka  

GSL1, Kiran, RTM 314, Kranti, Varuna

23-27

 

MS

 

11.

Leaf : growth habit (angle between leaf and horizontal)

erect(>85°)
semi erect (66-85°)  
open (46-65°)  
semi-prostrate (31-45°)  
prostrate (<31°)

1
3

5 
7  

9

 

23-27

VS

 

12.
(*)

Time of flowering (50% of the plant with at least one open flower)

very early (<31 days)
early (31-40 days) medium(41-50days)  
late (51-60 days)  


very late (>60 days)

1
3
5
7


9




Bhawani, PT303, TH68, Bio902  
GSL1, Pusa Gold, Kiran, Kiran, Varuna

 

MG

 

13.
(*)

Flower : colour of petals

white
cream
light yellow  


yellow
orange

1
2
3


4
5

 
RTM314
Bhawani, GSL1, Jhumka, Bio 902, Varuna
Kiran, PC5

62-63

VG

14.

Flower : length of petals

short (<1.2 cm)


medium (1.2-1.5 cm)  


long (>1.5 cm)

3


5


7

Bhawani, GSL1, Jhumka, Kiran, Bio902
PT303, Neelam, Pusa Gold, RH30, Rohini
GSL2

62-63

MS

15.

Flower : width of petals

narrow (<0.7 cm)  


medium (0.7-0.8 cm)


broad (>0.8 cm)

3


5


7

GSL1,Juumka, Kiran, Bio 902, Varuna
PT303, Neelam, Pusa Gold, GM1, Rohini
Bhawani, RTM314

62-63

MS

 

16.

Anther : production of pollen

absent  
present

1
9

 
Bhawani, GSL1, Jhumka, Kiran, Varuna

62-63

VS

17.
(*)
(+)

Plant : main shoot length

short (< 41 cm)

medium (41-60 cm)

long (>60 cm)  

3

5

7

GSL1, PusaGold, Kiran, RTM 314
Bhawani, GSL2, Jhumka, Varuna
GM1

75-80

MS

 

18.

(*)

Plant : height (including side branches)

rapeseed
short (<51 cm)  
medium (51-75 cm)
tall (76-100 cm)
very tall (>100 cm)
 

mustrard
short (<101 cm)  
medium(101-150 cm)  

tall (151-200 cm)

very tall (>200 cm)

 


3
5
7
9


3
5

7

9

 




PT507, PT303
Bhawani, JT1, TH68  



RH30, SEJ2, Kranti
Varuna, Rohini, Bio902

75-80

MS

 

19. (+)

Siliqua : length

short (<4.5 cm)


medium (4.5-5.5 cm)


long (>5.5 cm)

3


5


7

Bhawani, Pusa gold, Kiran RTM314,Varuna
TH68, Neelam, Jhumka, PC5, RH30
GSL1, GSL2, PBR91, Pusa Bold

75-80

MS

 

20. (+)

Siliqua : length of beak

short (<0.8 cm)  
medium (0.8-1.2 cm)

long (>1.2 cm)

3
5

7

PC5, RTM314
Bhawani, Kiran, Basanti, Rohini
PT303, Neelam, GSL1, Jhumka

75-80

MS

21.

Siliqua : number on main shoot

few (<30)


medium (30-50)


many (>50)

3


5


7

Pusa gold, Jhumka, Kiran, RTM314, RH30
Bhawani, GSL2, Bio902, Rohini, Varuna
GSL1, Sheetal

75-80

MS

 

22. (+)

Siliqua : density  on main shoot

 

low  
medium


high

3
5


7

Kiran, RTM314
Bhawani, Pusa Gold, Varuna, Bio902  
GSL1,KS101, Rohini

75-80

MS

 

23.

Siliqua : angle with main shoot/branches

appressed (< 21Ί)
semi-appressed (21-30Ί)
open (>30Ί)

3
5

7

 

 
GM1, Vardan  
Bhawani, GSL1, Pusa Gold, Kiran, RTM314, Basanti, Bio 902

75-80

MS

24. (+)

Siliqua : surface texture

smooth


intermediate

constricted

1


2

3

Bhawani, GSL1, JM1, Jhumka, RTM314
TH68, Sheetal, Bio902, Varuna

JT1, Kiran, Basanti

80-85

VS

 

25.

Siliqua : number of seeds per siliqua

few (<11)  
intermediate(11-20)


many (>20)

3
5


7

 
TH68, Sheetal, Kiran, RTM314, Varuna
GSL1, Pusa gold, PC5

80-85

MS

 

26. rapeseed
 early(<81 days)
medium(81-100days)
late(101-120days)

very late(>120days)

mustared
 early(<111 days)
medium(111-130days)
late(131-150days)

very late(>150days)


3
5
7

9


3
5
7

9




Bhawani, JT1, Jhumka PT303




Kiran, RH30, Varuna, BIO902

27.
(*)

 

Seed : colour

yellow
dull grey  
reddish brown  

brown

black

1
2
3

4

5

Jhumka, Basanti
Kiran, RTM314
TH68, GSL1, CS52
Bhawani, PC5, RH30
GSL2, Bio92, Varuna

00

VG

28.
(*)

Seed : size (weight of 1000 seeds)

rapeseed
 small  (<3.0 g)  
medium (3.0 - 4.0 g)  

large ( > 4.0 g)

mustard
small (< 4.0 g)  
medium (4.0-5.0 g)  
large (>5.0 g)


3
5

7


3
5
7



Bhawani,GSL1, RTM 314
GSL2, Jhumka  


Kranti
Kiran, JM1
Varuna, Bio 902, CS52, SEJ 2

00

MG

 

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VIII. Explanation on the Table of Characteristics

Characteristic 1. Seed : erucic acid

The erucic acid content should be observed on seed sent in by the applicant. It should be expressed as percentage and estimated with NUCON-5765 GLC using following specifications:

Column                       : Stainless steel SP 2300 (2%)+ SP 2310 (3%)

Temperature              : Oven (2250C), injector (2250C) and detector (2500C)

Gas flow                     : N2      -           40-50-ml/ cm2

                                      H       -           4-kg/ cm2

                                      Air     -           2-kg/ cm2

Reference                  : (i) Morision W.R. and Smith L.M. (1964): Preparation of Methyl
                                          esters. J. Lipid Res. 5:600

                                      (ii) Met Calfe L.D., Scmnitz A.A. and Pelke (1966): Anal Chem.
                                           38:514

 

Characteristic 2. Seed : glucosino

Glucosinolate content should be measured on seed sent in by the applicant. It should be estimated by complex formation between glucosinolate and tetrachloropaladate (II) using Micro scan ELISA Reader (MS 5605). Kumar et al.

 

Characteristic 3. Seed : oil content

Oil content of dry seed sent in by the applicant is determined using NIR & NMR (Non destructive techniques).

 

Characteristic 6. Leaf : lobes

Absence or presence of lobes should be observed on the whole plant at rosette stage. Parts of the leaf blade are considered as lobes if their length is at least equivalent to the width of the leaf petiole at their point of attachment and if the upper notch of the blade has at least half the length of the lobe itself. Secondary lobe(s) are not counted.

1
Absent		 2
Present

 

 Characteristic 7. Leaf : number of lobes

 In case lobes present, these are counted as shown below:

 

Characteristic 8-10.  Leaf: Dentation (8), length (9), width(10)

Leaf length, width, dentation and of petiole are measured as shown below:

7= Part on which the dentation should be recorded

 

Characteristics 17. Plant : main shoot length

Total length of the siliquaes-bearing portion (main raceam) of the main stem.

 

Characteristics 19 and 20. Siliqua : length (between pedicel and beak (19)), length of beak (20)

 

Characteristics 22. Siliqua : density on main stem

It should be computed as a ratio between main shoot length and number of silique borne on main shoot. 

 

Characteristics 24. Siliqua : surface texture 

1
smooth		 2
intimidate		 3
constricted

 

Characteristics 26. Maturity duration

Maturity duration should be recorded as days from date of sowing to when 75% of siliquae turn yellowish.  

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IX. Technical Questionnaire

 

Reference Number
(not to be filled in by the applicant)
 

Technical questionnaire  
to be completed in connection with an application for plant breeders' rights  

 
1. Species:

Yellow sarson (Brassica rapa var. yellow sarson), brown sarson (B.rapa var. brown sarson), toria (B.rapa var. toria), toria), Indian mustard (B. juncea L.), karanrai (B. carinata), gobhi B.napus L.) and black mustard (B.nigra L.)

 
2.      Applicant (Name and address) :  

 

 

 

 
3.  Proposed denomination or breeder's reference

  

 

  

4.  Information on origin, maintenance and reproduction of the variety

 

4.1 Type of material

(a)   inbred line
                       - male sterile line ( )
                       - male fertile line ( )
(b)   hybrid
                         - male sterile hybrid ( )
                      - male fertile hybrid ( )
(c)   open pollinated variety
 
(d)   synthetic variety
  
(e)   other (please indicate)
 
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4.2  Formula (if applicable, for each component in separate sheets, the information
according  to the following chapter 5 to 7 to be added

Single hybrid

-Denomination or breeder’s reference
  of female parental line                                           …………………………

 -Denomination or breeder’s reference 
  of male parental line                                               …………………………

 

Three-way hybrid

  Denomination or breeder reference of :

- Single hybrid used                                                   ………………………………

- female parental line of the single hybrid                  ………………………………

      - male parental line of the single hybrid                     ………………………………

- female parental line of the three-way hybrid            ………………………………

- male parental line of the three-way hybrid               ………………………………
        

NB: In case of use of male sterility system, indicate the name of the maintainer line of the female parental line                                                   ……………………………

In case of use of self-incompatibility system, indicate if applicable the name of the self-compatible line                                                              ……………………………


4.3  Genetic and breeding method 
  
 

4.4  Other Information

 

 

5. Characteristics of the variety to be indicated (the number in brackets refers to the corresponding characteristic in Test Guidelines; please mark the state of expression which best corresponds).  
S.No. Characteristics States of expression Example varieties  Notes

5.1
(6)

 Leaf : lobes

absent
present

   

1 [ ]
9 [ ]

5.2
(12)

 Time of flowering

very early
early
medium
late
very late

   

1 [ ]
3 [ ]
5 [ ]
7 [ ]
9 [ ]
5.3
(16)		 Anther : production of pollen

absent
present

  

1 [ ]
9 [ ]

5.4.
(17)

Plant : main shoot length

short
medium
long

   

3 [ ]
5 [ ]
7 [ ]
6. Similar varieties and differences from these varieties  

Denomination of
Similar variety

Characteristic in
which the similar variety is different 0)

State of expression of similar Variety

 State of expression of candidate variety
                          
0)  In the case of identical states of expressions of both varieties, please indicate the size of the difference.  

7. Additional information which may help to distinguish the variety

 

      7.1 Resistance to pests and diseases  

          

 

      7.2 Special conditions for the examination of the variety, if any

 

      
        7.3 Main use

a)     Oil

b)     Forage

c)      Other (specify)

        7.4 Other information

 

8.  Authorization for release

 

(a)   Does the variety require prior authorization for release under legislation concerning the protection of the environment, human and animal health?

 

                     Yes              {     }                                                         No         {      }

 

(b)    Has such authorization been obtained?

 

                      Yes             {     }                                                         No        {      }     


If the answer to that question is ‘Yes’, please attach a copy of such an authorization   

 

[End of document]

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