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R
elationship SSR-Based Molecular Marker Cotton F
1
Inter Specific Hybrids Performance Seed Cotton Yield
28
flowers were pollinated during the next day morning
between 9.30 am to 11.30 am by brushing the pollen
from one of the F
4
lines (used as male) on the
stigmatic surface. The pedicel of each pollinated
flower was tied with price label bearing date and lines
number for identification of crossed bolls. In this
manner derived F
1
s seeds were obtained.
Simultaneously, two populations of F
4
lines were
selfed and material was advanced to F
5
generation
during the same season.
Table 5 List of F
4
barbadense line parents involved in the study
Sl.No
Abbreviation
Lines
1
L1
(DB 533 × DB 534 F
5
IPS 44)
2
L2
(DB 533 × DB 534 F
5
IPS 62)
3
L3
(DB 533 × DB 534 F
5
IPS 105)
4
L4
(DB 533 × DB 534 F
5
IPS 26)
5
L5
(DB 533 × DB 534 F
5
IPS 71)
6
L6
(DB 533 × DB 534 F
5
IPS 30)
7
L7
(DB 533 × DB 534 F
5
IPS 25)
8
L8
(DB 533 × DB 534 F
5
IPS 49)
9
L9
(DB 533 × DB 534 F
5
IPS 23)
10
L10
(DB 533 × DB 534 F
5
IPS 36)
11
L11
(DB 533 × DB 534 F
5
IPS 15)
12
L12
(DB 533 × DB 534 F
5
IPS 1)
13
L13
(DB 533 × DB 534 F
5
IPS 33)
14
L14
(DB 533 × DB 534 F
5
IPS 24)
15
L15
(DB 533 × DB 534 F
5
IPS 16)
16
L16
(DB 533 × DB 534 F
5
IPS 52)
17
L17
(DB 533 × DB 534 F
5
IPS 12)
18
L18
(DB 534 × DB 533 F
5
IPS 22)
19
L19
(DB 533 × DB 534 F
5
IPS 14)
20
L20
(DB 533 × DB 534 F
5
IPS 34)
21
L21
(DB 533 × DB 534 F
5
IPS 55)
22
L22
(DB 533 × DB 534 F
5
IPS 17)
23
L23
(DB 533 × DB 534 F
5
IPS 32)
24
L24
(DB 533 × DB 534 F
5
IPS 38)
25
L25
(DB 533 × DB 534 F
5
IPS 48)
26
L26
(DB 533 × DB 534 F
5
IPS 13)
27
L27
(DB 533 × DB 534 F
5
IPS 6)
28
L28
(DB 533 × DB 534 F
5
IPS 8)
The experimental material was planted on a medium
black soil at College of Agriculture, Dharwad under
irrigated condition. The F
5
lines, derived F
1
s of two
populations along with the straight crosses and ruling
commercial check (MRC6918 and DCH 32) were
sown during kharif 2011 in all a randomized block
design with two replications and a spacing of 90 cm
between rows and 60 cm between the plants within a
row. Recommended fertilizer doses were applied and
other cultural practices were carried out at regular
interval. Plant protection measures were taken at
appropriate time to control pests and diseases. Each
set of 28 F
4
lines thus was involved in 112 crosses
(refer to as derived F
1
s), which were subjected to L×T
analysis. The observations were recorded for number
of bolls (no/plant), mean boll weight (g), seed index
(g), ginning outturn (%), lint index (g), seed cotton
yield (kg/ha), fiber length (mm), fiber strength (g/t),
fiber micronaire value (µg/inch), fiber uniformity ratio
(%), fiber maturity ratio and fiber elongation (%).
Fiber quality traits were measured with the High-
Volume Instrument.
2.2 SSR molecular marker analysis
Leaf tissue of each parents was harvested and total
genomic DNA was extracted from young leaves using
the hexadecyl-trimethyl ammonium bromide (CTAB)
method described by Saghai-Maroofet al., (1984).
SSR assays were performed using 40 oligonucleotide
primers from Sigma Aldrich Chemicals Pvt. Ltd., Co.
Amplification reactions were carried out in 20 mL
volumes containing 2.0 mL 10× assay buffer, 2.0 mL
dNTP mix (2.5 mM each), 0.5 mL forward primer
(5 pM/mL), 0.5 ml Reverse (5 pM/mL), 0.5 mL
Taq
DNA polymerase (3U/mL), 2.0 mL Template DNA
(15 ng/mL) and 7.5 mL Sterile double distill water.
The amplification programmed for 5 min at 94
℃
Denaturation (initial) of genomic DNA by one cycle
followed by 25 cycles of 1 min at 94 , 1 min at 48 ±
℃
5 and 1 min at 72 . This was followed by a final
℃
℃
extension at 72 for 5 min. Amplification products
℃
were analysed by Non-Denaturing gel electrophoresis
(PAGE) and viewed by silver staining.
2.3 Scoring the amplified fragments
The amplification of DNA profiles for all the primers
were compared with each other and the bands of DNA
at each amplification level of every primer were
scored as present (1) or absent (0) thus generating the
0, 1 matrix.
Per cent polymorphism (%) = (Total No. of
polymorphic bands)/(Total No. of bands generated by
40 primers) ×100%
Genomics and Applied Biology