MSc

Cocoa (Theobroma cacao L.) also known as ‘chocolate tree’ is a major cash crop of
tropical countries and belongs to the family Malvaceae. The plant is a native of Andes, South
America and was introduced to India during 1970s. In India, cocoa is cultivated as an important
intercrop with in coconut, arecanut, rubber, oil palm etc. The statistics shows that, 80 per cent
of cocoa plantations in India is established with planting materials distributed from Kerala
Agricultural University (KAU).
In India, the genetic base of cocoa is widened by the systematic introduction of
germplasm from University of Reading, UK. Cocoa Research Centre (CRC), KAU holds
Asia’s largest germplasm with 640 accessions. Exploitation of these germplasm has resulted in
the release of 15 cocoa varieties from KAU.
Central sub-committee on crop standards, notification and release of varieties for
agricultural crops has made it mandatory to provide DNA finger printing data along with the
varieties where ever the proposal for national release/ notification is submitted. DNA markers,
that are not subjected to environmental influences act as an efficient tool to identify and
differentiate accessions and cultivars which are similar in morphological characteristics and
with indistinct traits. DNA finger printing is successfully applied for cultivar identification,
controlling seed purity of hybrids and checking the genetic similarity between cultivars. Hence,
the technique act as a powerful tool to protect Plant Breeder’s Right (PBR).
In the present investigation, eight promising cocoa varieties CCRP 1, CCRP 2, CCRP
4, CCRP 5, CCRP 6, CCRP 7 (selections), and CCRP 8 and CCRP 9 (hybrids) released from
KAU were characterized using morphological and molecular markers. For morphological
characterization, six qualitative and nine quantitative characters were recorded. And it was
observed that, CCRP 6 and CCRP 8 were superior based on the performance of their yield
contributing characters.
Molecular characterization was performed with genomic DNA isolated using modified
Chandrakant’s (2014) protocol. Based on the polymorphism, ten ISSR (inter simple sequence
repeats) and eleven SSR (simple sequence repeats) primers were selected. These selected
primers were used for developing DNA fingerprints of varieties under study. In the further
analysis, amplicon generation pattern were carefully scored to locate polymorphism. IndividualISSR and SSR amplification pattern further converted into variety wise fingerprints and thus
consolidated DNA fingerprints on each marker system were developed.
ISSR primers UBC 810 and UBC 826 were found to differentiate CCRP 6 from other
genotypes. Primers UBC 827, UBC 846 and UBC 866 were generated unique amplicons in
CCRP 9. UBC 841 and UBC 846 were capable of distinguishing CCRP 5 from other genotypes.
Primers UBC 835 and UBC 866 were generated amplicons in hybrids CCRP 8 and CCRP 9
alone and the markers can be used for differentiating these hybrids.
SSR marker analysis was performed using selected eleven primers. Selected primers
generated polymorphic amplicons and were capable of distinguishing between varieties.
Primer mTcCIR 8 generated unique amplicon at 220 and 420 base pairs (bp) in CCRP 1 and
CCRP 4 respectively which was a fingerprint for those varieties. The unique amplicon
generated by mTcCIR 33 at 320 bp was a fingerprint of CCRP 2. Polymorphic amplicons
generated by mTcCIR 42 at 200 bp (CCRP 4) and at 220 bp (CCRP 9) were fingerprints of
respective varieties.
In future, more morphological characters have to be screened and correlated with the
markers shared by two or three varieties. This will help to know whether the shared bands are
responsible for expression of some distinct traits. DNA fingerprints for remaining seven
released cocoa varieties also have to be designed.