Use of Response Surface Methodology (RSM) to Optimize Culture Media for in vitro Callogenesis in Ipomoea obscura (L.) Ker-Gawl, a Little Known Medicinal Plant

Aims: The present study was undertaken to explore the potentiality of in vitro micropropagation (by
callus induction) of Ipomoea obscura(L.) Ker-Gawl (Convolvulaceae), a perennial herb.
Method: Axenically grown cotyledonary leaves regenerated into profuse calli, in various combinations
of naphthaleneacetic acid (NAA), indole-3-acetic acid (IAA), indole-3-butyric acid (IBA) and 2,4-
dichlorophenoxyacetic acid (2,4 D) with benzylaminopurine (BAP). Microroots emerged from these
calli when subjected to various combinations of NAA and BAP. Maximum number of root meristems
and microroots were formed in 1.25 mg/L NAA with the combination of 0.5 mg/L BAP (1:4 v/v). Leaf,
nodal and internodal segments as explants were cultured on MS media containing 2,4 D at 0.5, 1.25,
2.50, 5.0 and 10.0 mg/L, for callus induction. Only leaf tips initiated callogenesis with highest
response at 10 mg/l 2,4 D and lowest with 0.5 mg/l 2.4 D. This is the first report of in vitro response of
I. obscura.
Results: All the results were analyzed on the context of response surface methodology (RSM) and
simultaneously by contour plot. Two variables lay out in X and Y axes and a third variable Z is plotted
for contour level as concentric values. An efficient and effective micropopagation protocol was
developed from the leaf of Ipomoea obscura (L.) Ker-gawl, an important medicinal plant.
Transplantation of regenerated plants under in vivo condition has been done successfully. RSM has
been used for modeling, analyzing and optimizing in vitro responses of Ipomoea obscura. The specific
target response surface plot and contour plot are focused to determine optimum shoot and root length
against optimum hormone concentration, number of shoot and root length in best concentration of
MS medium and finally mean number of root and shoot in optimum NAA concentration. Depending
upon variances of dry weight of callus, rooting response and root length, response surface plots
(RSM) and contour plots were drawn where four types of hormone combinations (IAA- BAP, IBABAP,
NAA-BAP, 2,4D-BAP) were used in different combinations.