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IN VITRO PROPAGATION OF BANANA (Musa spp.)
(Department of Biotechnology, Sher-e-Bangla Agricultural University, 2013-12) KHATUN, FAHIMA
The experiment was conducted at the Biotechnology Laboratory, Department of Biotechnology. Sher-e-Bangla Agricultural University. Dhaka, during October 2013 to August 2014 to investigate effects of hcnzylaminopurine (RAP) concentrations (1.0. 10, 3.0. 4.0 and 5.0 mg/L) and indole-3-butyric acid (IDA) (0.5. 1.0. 1.5. 2.0 and 2.5 mg/L) for in vitro regeneration of Sagar and Sabri Banana varieties using shoot tip explants. The highest response of explants 88% in Sagar and 84% in Sabri were with 4.0 and 5.0 mgfL BAP. whereas in 4.0 mg/L BAP+2.0 mg/L IRA had 85% response in Sagar and 5.0 mg/L BAP+2.5 mg/L IBA had 90% response in Sabri. In both Sagar and Sabri. minimum number of days for shoot initiation (7.80 and 9.40 days), maximum number of shoots per explant (6.80 and 3.40), highest shoot length (8.31 and 6.19 cm), number of leaf (4.24 and 4.49) were observed in 4.0 mg/L and 5.0 mg/L RAP. In contrast, due to combined effect, the highest shoot number per explant (5.60 and 3.40) was in 4.0 mg/L BAP+1 .5 mg/I. IRA and 5.0 mg/L I3AP+2.0 mg/L IDA in Sagar and Sabri. Again, root formation required minimum 7.60 and 8.00 days in 1.5 mg/L IRA for Sagar and Sabri, respectively. The highest number of roots (5.20 per explant) was observed in 0.5 mg/L of IRA for Sagar and that for Sabri was 3.40 per explants observed in 1.5 mg/L of IRA at 6 WAI. In combination both for Sagar and Sabri, the highest number of roots (5.00 and 5.20 per explant) and root length (6.61 and 6.72 cm) were produced by 3.0 mg/L BAP±1.0 mg/I. [BA. In natural environment. Sagar variety survived more (94.74%) in soil than Sabri variety (88.89%). Ultimately combined effect of RAP and IRA seemed better than individuals based on average performance of growth parameters and the protocol optimized in the present study could be used in vitro rapid propagation of Banana plantlets. iv
IN VITRO PLANT REGENERATION OF Aloe vera L.
(Department of Biotechnology, Sher-e-Bangla Agricultural University, 2014-07) ALAUDDIN, MD.
The present research was carried out on Aloe vera L. in Biotechnology Laboratory of the Department of Biotechnology, Sher-e-Bangla Agricultural University, Sher-e- Bangla Nagar, Dhaka-1207 during the period from September, 2013 to July, 2014 to evaluate the effect of different plant growth regulators on multiple shoot proliferation and root formation along with acclimatization for in vivo survival and finally to develop a viable micropropagation protocol. Shoot tip of young lateral shoots (sucker) were used as the explants, which were sterilized using freshly prepared 0.1% HgCl2 solution with few drops of Tween-20. After sterilization, explants were inoculated in MS media supplemented with either singly or in combination of the Benzylamino purine (BAP), Kinetine (KIN), Indolebutyric acid (IBA), Naphthaleneacetic acid (NAA). The highest percentage of shoot proliferation (85%) and minimum days to shoot induction (13.6 days) were achieved on MS medium containing 1.0 mg/L BAP + 0.5 mg/L IBA. The highest 14.8 shoots and 24.4 leaves per explant were also observed in the same media composition. But the highest average length of shoots (9.62 cm) was recorded from media containing 1.5 mg/L BAP + 1.0 mg/L NAA. The highest percentage (95%) of rooting was obtained in media treated with 3.0 mg/L IBA along with maximum 12.4 roots per explant. But the highest (10.24 cm) average length of root was observed in 2.0 mg/L IBA. The highest (5.374 mm) diameter of root and minimum days to root induction (10.6 days) were noticed on the same culture media supplemented with 2.5 mg/L NAA. In regenerated plantlets, 100% survival in growth chamber conditions and 88% survival under shade house and 90% survival in open atmosphere were achieved. Micropropagation was found to be very effective and promising method in the propagation of Aloe vera and this protocol can be a useful tool for proliferation of Aloe vera.
ASSESSMENT OF STRESS-TOLERANCE ATTRIBUTES IN WHEAT USING GENE – SPECIFIC MOLECULAR MARKERS
(Department of Biotechnology, 2014-12) MAHMUD, RONA
DNA PROFILING AND MOLECULAR DIVERSITY ANALYSIS OF BRINJAL (Solanum melongena L.) CULTIVARS USING RAPD MARKERS
(Department of Biotechnology ,Sher-e-Bangla Agricultural University , Dhaka 1207, 2014-12) SULTANA, SHARMIN
An experiment was conducted on Brinjal (Solanum melongena L.) in Biotechnology Laboratory of the Department of Biotechnology, Sher-e-Bangla Agricultural University, Dhaka-1207. The genetic variation of 11 brinjal varieties (BARI begun 1, BARI begun 4, BARI begun 5, BARI begun 6, BARI begun 7, BARI begun 8, BARI begun 9, BARI begun 10, BARI hybrid begun 1, BARI hybrid begun 3 and BARI hybrid begun 4) were studied based on PCR technology using five RAPD primers. Sum total of 44 distinct DNA amplified band were observed with an average of 5.6 polymorphic bands per primer. The overall gene diversity was detected 0.2186 and level of polymorphism was detected 63.64%. The pair wise inter-variety similarity indices ranged from 67.03% to 97.61%. The UPGMA Dendrogram segregated 11 genotypes of brinjal into two main clusters. The first major cluster had only one genotype (BARI begun 6) and the second major cluster had rest of ten genotypes. BARI begun 6 vs. BARI begun 1 showed highest Nei’s genetic distance (0.5261) as they are released from different parental origin. On the other hand BARI began 9 vs. BARI begun 7 varietal pair showed lowest genetic distance (0.0465) as they are released from same parental origin. The experiment reveals significant level of variations among 11 brinjal genotypes by RAPD markers. The results of this study can be used as a baseline for the future diversity assessment and genetic analysis of brinjal varieties of Bangladesh.
IN VITRO REGENERATION OF Caladium bicolor
(Department of Biotechnology, Sher-e-Bangla Agricultural University, 2014-12) AHMED, KHANDAKER SOHAEL
The experiment was conducted at the Biotechnology Laboratory, Department of Biotechnology, Sher-e-Bangla Agricultural University, Dhaka, Bangladesh during the period from October 2013 to July 2014 to investigate the effects of BA, Kin, IBA, IAA, NAA for in vitro regeneration of Caladium bicolor using shoot tip explants. The work was designed in CRD with three replications. Maximum number of shoot (5) initiated when MS media was supplemented with 1.0 mg/L BA and minimum (1) was at control. Highest percentage (90%) of explants showed shoot induction at BA 0.50 mg/L +2.00 mg/L IBA and minimum shoot induction days (40.47) were recorded at BA 1.00 mg/L +2.00 mg/L IBA. Highest number of shoots (19.67) were recorded at BA 1.00 mg/L +2.00 mg/L IBA. Longest length of shoot (11.43 cm) was recorded at BA 0.25 mg/L +2.50 mg/L IBA. Maximum number of leaves were recorded at BA 1.00 mg/L +2.00 mg/L IBA. To investigate root formation potentiality and root morphology NAA, IAA and IBA at different concentrations were used. Among the rooting hormones IAA at 2.0 mg/L concentration showed maximum percentage (90%) of root induction. Maximum numbers (29.87) of root were recorded when IAA was used at 2.0 mg/L concentration. Highest length of root (10.20 cm) was recorded at 2.0 mg/L IAA. Ultimately combined effect of BA and IBA seemed better than individuals based on average performance of growth parameters. Survival rate was 92% in open field condition. Reliable protocols for micropropagation of Caladium bicolor were established and could be used for large scale production of disease free, high yielding and premium quality planting material.
IN VITRO PROPAGATION OF GINGER (Zingiber officinale
(Department of Biotechnology, Sher-e-Bangla Agricultural University , Dhaka 1207, 2014-12) FERDOUS, RUMANA
The present research was carried out in Biotechnology Laboratory of the Department of Biotechnology, Sher-e-Bangla Agricultural University, Dhaka-1207 from the period of September 2013 to July 2014 to evaluate the effects of different plant growth regulators on in vitro propagation of ginger (Zingiber officinale). The healthy, disease free sprout was used as explants. Explants was inoculated in MS (Murashige and Skoog), media supplemented with the Kinetine (KIN), alone and in combination with GA3 and Indolebutyricacid (IBA). The highest percent of shoot induction (90%) was observed on 3.0mg/L+1.5mg/L GA3 and the minimum days to shoot induction (13.00 days) were achieved on KIN 3.0mg/L.The highest number of shoot (4.6), maximum number of leaf per explants (10.00) and average length of shoot (4.56 cm) were recorded in 3.0 mg/L KIN+ 1.5 mg/L GA3.The highest percent of root induction 95% and the maximum (14.2) cm length of root induction were noticed on 2.5 mg/L IBA. Micropropagation was found very effective and promising method in the proliferation of ginger and can be used for large scale production of disease free and quality planting material.
INVESTIGATION OF GLYOXALASES IN Brassica spp UNDER ABIOTIC STRESS AND PURIFICATION OF GLYOXALASE-I
(Department of Biotechnology, Sher-e-Bangla Agricultural University, 2014-12) HOSSAIN, MD. SHAHADAT
This study was carried out in the Molecular Breeding Laboratory of Bangladesh Agricultural Research Institute (BARI), Joydebpur, Gazipur during the period from March, 2014 to March, 2015 to investigate the role of Glyoxalase system in contrast plants of Brassica spp. Ten prescreened genotypes were subjected to test the variation among them with 5 known SSR markers, and among them, E5 (Tori 7) and E6 (BARI sarisha 16) were selected based on their variation. The comparative phenotyping study of E5 and E6 under salinity and drought stress revealed that E6 is more tolerant than E5 in both stresses. To investigate the responses of Glyoxalase-I (Gly-I) and Glyoxalase-II (Gly-II) in detoxification of methylglyoxal (MG) under salinity and drought stress, 5 day old Brassica seedlings were subjected to impose NaCl-induced salinity level (16dS/m) and drought stress and data were observed at 2 day, 4 day and 6 day of stress. The contents of (MG) increased 2.0, 2.5 and 3.0 fold over control under salinity at 2, 4 and 6 day, respectively in E6 and 0.5, 0.3 and 0.6 fold in E5. On the other hand, in drought stress, methylglyoxal (MG) level increased 2, 3 and 3.5 fold over control at 2, 4 and 6 day, respectively in E6 and 2, 2.5 and 3 fold in E5. Under salinity stress, Gly-I activity increased by 41%, 69% and 105% in E6, at 2, 4 and 6 day, respectively, whereas 12%, 43% and 41% in E5. On the contrary, in drought stress, Gly-I activity increased by 70%, 76% and 61% over control at 2, 4 and 6 day, respectively in E6, and 37%, 32% and 53% in E5. In E6, Gly-II activity increased 144%, 128% and 101% at 2, 4 and 6 day of salinity stress, respectively in E6, and 52%, 59% and 50% in E5. Notably, concomitant increased activities of Gly-I and Gly-II with increased MG suggested their MG detoxification role in Brassica Spp. At the same time, it was remarkable that both Gly-I and Gly-II activities were higher in E6, and hence, Gly-I was purified from E6 seedlings. In purification, Gly-I were found to be highly purified and migrated as a single band on SDS-PAGE with an apparent molecular mass of 27 kDa in silver staining and coumassie brilliant blue staining with purification fold of 113 and recovery of 0.38%. iii
MOLECULAR DIVERSITY ANALYSIS OF SOMACLONAL VARIANTS IN POTATO (Solanum tuberosum L.) BY RAPD MARKERMD SHAMSUZZAMAN
(Department of Biotechnology, Sher-e-Bangla Agricultural University , Dhaka 1207, 2014-12) SHAMSUZZAMAN, MD
An experiment was carried out in Biotechnology Laboratory of the Department of Biotechnology, Sher-e-Bangla Agricultural University (SAU), Dhaka, Bangladesh to study the DNA fingerprinting and genetic diversity of nine somaclonal variants of potato (Solanum tuberosum L.) and three check varieties (Cardinal, Diamant and Asterix). The somaclonal variants were SIP-3, SIP-5, SVP-6, SVP-18, SVP-19, SVP-25, SVP-55, SVP-56 and SVP-68. Genomic DNA was extracted from young leaves and PCR reaction was performed with six RAPD primers. Sum total of 54 distinct and differential amplified DNA bands were generated from PCR amplification. Out of them, 47 bands (87.04%) were polymorphic and 7 bands (12.96%) were monomorphic. The pair wise inter-variety/genotype similarity indices ranged from 61.59% to 93.55% with an average of 74.31%. The Nei’s genetic identity among 12 potato genotypes ranged from 0.5370 to 0.9074 and genetic distance ranged from 0.0972 to 0.6217. The UPGMA Dendrogram segregated the 12 potato genotypes into two main clusters. The first cluster contained 8 genotypes and the second cluster had 4 genotypes. The Dendrogram also indicated the highest genetic distance and the lowest genetic identity between SVP-6 vs SVP-68 genotype pair. The study revealed a significant amount of relationship and genetic diversity among the studied 12 potato genotypes. The RAPD markers were found to be useful tool for DNA fingerprinting, polymorphic study and genetic diversity analysis of potato genotypes with a great potential for the Bio technologists and Plant breeders for the improvement of potato cultivars
CALLUS INDUCTION AND MICROPROPAGATION OF PINEAPPLE (Ananus comocus L.)
(Department of Biotechnology, Sher-e-Bangla Agricultural University, 2015-12) AKTER, FATEMA
The present study was undertaken in Biotechnology Laboratory of the Department of Biotechnology, Sher-e-Bangla Agricultural University, Sher-e-Bangla Nagar, Dhaka 1207 during the period from January. 2014 to December. 2014 to evaluate the effect of different plant growth regulators on callus induction, shoot proliferation, root formation and to develop micropropagation protocol of pineapple. Crown tip of pineapple fruits were used as explants and inoculated in MS media supplemented with different cone. of Benzyl adenine (BA), Kinetine (KIN). Indolebutyric acid (IBA). Naphthaleneaeetie acid (NAA) alone or in combination with each other. The treatment 3.0 mgIL KIN + 2.5 mg/L NAA showed the highest percentage (800/0) of callus induction. But minimum days (21.20) to callus induction and maximum weight of callus (2.80 gm) was observed with 2.0 mg/L KIN + 1.5 mg/L NAA treatment. The highest 85% shoot induction was obtained with 3.0 mgfL KIN +1.0 mg/L NAA treatment and minimum 7.60 days was recorded with the same treatment. The highest number of shoots (18.40) after 6 weeks of subculture was recorded in 3.0 mg/L KIN + 1.0 mg/L IBA and the longest shoot (9.04 cm) was found with 3.0 mg/L KIN+2.0 mg/L IBA. The treatment 2.0 mg/L IBA+ 1.5 mg/L NAA showed the highest percentage (75%) of root induction, minimum 16 days to root induction and the longest root (7 cm). But the highest number of roots (15.60) per explants was recorded with 3.0 mg/L IBA+ 1.0 mg/L NAA after 6 weeks of subculture to rooting media. Regenerated plantlets showed, 85% survival in growth chamber conditions and 82% in shade house stage of hardening with Sand : Soil : Cow-dung (1:1:1) and 78 % in open atmosphere at direct sunlight.
IN VITRO PROPAGATION OF BETEL VINE
(Department of Biotechnology, Sher-e-Bangla Agricultural University, 2015-12) ELAIH, MD. MAHABUB
The experiment was carried out on Betel vine (Piper belle L.) in Biotechnology Lab. of the Department of Biotechnology, Sher-e-Bangla Agricultural University, Sher-e Bangla Nagar, Dhaka-1 207 from the period of July 2014 to April 2015 to investigate the effect of BA, KIN, IAA and IDA on shoot proliferation, root formation and micropropagation protocol development. Nodal segment was used as explant. The highest percent (70%) of shoot proliferation and minimum days to shoot induction (21.67 days) were achieved on MS medium containing BA 1.0 mg/L + KIN 3.0 mg/L. Maximum average length of shoot (8.43 cm) was reported from media containing BA 0.5mg/L + KIN 4.0 mg/L. The highest percent (90%) of rooting and maximum 14.40 root/explant were reported in media treated with 3.0 mg/L IRA. Maximum 90% survival was observed in growth chamber and 83% during shade house and 64% in open atmosphere. A protocol for micropropagation of Betel vine, has been established and this experiment can be a useful tool for micropropagation of Betel vine.
MORPHOLOGICAL CHARACTERIZATION OF POTATO VARIETIES UNDER IN VITRO AND IN VIVO CONDITION
(Department of Biotechnology, Sher-e-Bangla Agricultural University, 2015-12) AKTER, KHADIJA
The experiment was conducted at the Biotechnology l.aboratorv and research farm of Sher-e-Bangla Agricultural University (SAU), Dhaka during the period from April 2015 to March 2016 to study the morphological characterizalior1 of potato varieties under in "i/ru and in nyu condition. The sprouts ol potato varieties were used as explants and 4 times sub-culture were done to study the morphological parameter of plantlet. The highest length of shoot (5.19 ciii) was in the variety Diamant and it was lowest in Asterix variety. The maximum diameter of shoot (1.16 mm) was recorded from Asterix, while the minimum diameter of shoot (0.82 mm) from Diamant. Due to the interac(ion effect of difiërent varieties and number of explants. the highest length of shoot (6.00 cm) was found in Diainant variety' when 8 explants used in each culture. while the lowest length (4.10 cm) in Cardinal variety used 3 explant per vial. In vitro regenerated plantlets were acclimatized under in vivo field condition. The variety Diamant showed the best performance in respect of all the parameters under this study. The weight of minituher and yield per plant were highest in Dianmnt (10.23 gm and 114.16 gni) variety whereas the lowest in Asterix (8.15 gui and 64.46 gm) variety. (uba y: I-::
CALLUS INDUCTION AND PLANT REGENERATION OF STEVIA (Stevia rehaudiana B.)
(Department of Biotechnology, Sher-e-Bangla Agricultural University, 2015-12) FERDOUS, JANNATUL
The present research work was carried out in Biotechnology laboratory of the Department of Biotechnology. Sher-e-Bangla Agricultural University. Shcr-e-Bangla Nagar, Dhaka- 1207 from the period of January 2014 to December 2014 to evaluate the effects of different plant growth regulators in in vitro on shoot proliferation and root lbrmation and finally to develop a viable micropropagation protocol in Stevia (Stevia rehaudiana) using shoot tip and nodal segments as explants. Explants was inoculated in MS (Murashige and Skoog) media supplemented with the Benzvladenine (BA) and Kinetine (KIN) for direct shoot formation. Combination of BA and Indolebutyricacid (IBA) as well as combination of KIN and IBA were used in MS medium for indirect shoot formation via callus induction. The highest percent of shoot induction (83.42%) obtained on 2.0 mg/L BA and the minimum days to shoot induction (10.6) were also obtained on 2.0 mg/I.. BA. The highest 10.8 shoots and 17.2 leaves per explants also observed in BA 2.Omg/L. But maximum average length of shoots per explants (3.94 cm) were reported from media containing 1mg/I. KIN. Highest callus induction potentiality (83.58%) was observed in 2 mg/L BA+1.5 mg/L IBA The minimum days to callus induction (13.4) were recorded in 1.0 mg/L KIN+I.5 mg/L IBA but the minimum days to shoot induction (9.6) from callus were recorded in 2.0 mg/L BA+1.5 mg/L IBA. Righest percent of root induction (78.18%) with 7.2 roots per explants and minimum 12.6 days to root induction were noticed on 1.0 mg/I. IBA. The combined ellèct on percent of' root induction was highest (76.92%) in the treatment of 1.0 mg/L KIN t 0.5 mg/L IBA. The same treatment showed minimum days (13.4) to root induction and maximum number of roots per explants. In acclimatization 90%, 80% and 80% survival rate were recorded in growth cabinet, shade house and field condition respectively. In vitro regeneration technique was found to be very effective and promising niethod in the proliferation of Stevia and this method can be used 11w large scale production of disease free and quality planting material.
EFFECT OF GIBBERELLIC ACID (GAs) ON GROWTH AND DEVELOPMENT OF RED AMARANTH (Amarnathus so.)
(Department of Biotechnology, Sher-e-Bangla Agricultural University, 2015-12) HYDER, K. M. AHSAN
An experiment was conducted to Jind out the efiets of diffettnt levels of GA Ofl tile 2rowtIl and yield of red amaranth at research farm. Sher-c-l3aitg!a Agricultural 1.Jniversity. l)llaka.during the period from October. 2014 to January. 201 6.It revealed that. 500 ppm 01 6A3 was best for lengihof stell) (20.27 cm). stein diamcteii 10.1 mm). total nuniherol leaf (28.58).length of edible te45.850cm). breath 01 edible leaIl2.983cm ). and ;tight of ten plants( 104.7gm). Ihe experiment also slnns that. maxnittni number of edible leaf was the highest 16.53. 21.04 and 24.12 in 12. 17 and 22 DASrespeetivelv in the 400 ppm GA3 treatment. Role of C tA on growth and development in red amaranth had also been studied under pot culture condition. Ii noticed that.500 ppm GA3 gave tile highest steni length 32.1 8 emho ever control treatment gave the lowest 26.18 cmrcsult at 12. 17 and 22 DAS. ?vlaxitnuut number of edible leaf 30.01 at 22 DAS was highest in 400 ppm (IA;. However control of CA3 uave the mini mm (17.26) result.CiA3 with 500 ppm exhibited niaxinium weight 60.33u with maximum gain of weight (84.42%) agallst 100 ppiii which showed titinimtim weight gain (19.271),n). The highest stein length growth ratcl .193. stein diatiieter growth ratc0.4633 and stem weight growth rate 2893 were obtLIiIlel with 500 ppm of CiA1 treatment. In pot culturecondition treatment 500 ppm C i.' ;respwidcd excellent in all the parameters under stud.
IN VITRO PROPAGATION AND NUCLEUS SEED PRODUCTION EFFICIENCY OF BARI DEVELOPED CLONE POTATO VARIETIES
(Department of Biotechnology, Sher-e-Bangla Agricultural University, 2016-12) SULTANA, NAZNIN
An experiment was conducted at Tissue Culture Lab and Net House of Tuber Crops Research Centre, Bangladesh Agricultural Research Institute (BARI), Joydebpur, Gazipur from April 2016 to February 2017 to investigate In vitro propagation and nucleus seed production efficiency of BARI developed clone potato varieties. In vitro shoots of potato varieties BARI Alu-35, BARI Alu-36, BARI Alu-37, BARI Alu-40, BARI Alu-46 and BARI Alu-62 were used as planting materials in this experiment. BARI Alu-7 was used as check variety. Significant difference were found among the growth parameters of in vitro plantlets of the potato varieties. The maximum number of leaves (14.80), length of shoot (99.98cm), weight of fresh shoot (1.66 mg) were observed in BARI Alu-36. Length of root of plantlets varied significantly among the varieties. The highest length of root (5.54 cm) was recorded in BARI Alu-36. The maximum number of primary root (17.00), number of secondary root (28.20) and fresh weight of root (0.359 mg) were observed in BARI Alu-36. BARI Alu-36 showed better performance on most of the morphological traits under in vitro condition. On the other hand, survival percentage and nucleus seed (G generation) production of potato have been calculated from virus free in vitro plantlets of potato at net house of TCRC, BARI during November 2016 to February 2017. Number of G seed tuber per plant 21.2, 16.0, 18.6, 19.4, 30.4, 37.4 and 52.4 were recorded in BARI Alu -7, BARI Alu -36, BARI Alu-37, BARI Alu-62, BARI Alu-40, BARI Alu-35, and BARI Alu-46 respectively. The maximum weight of G 1 1 seeds per plant were found from the variety BARI Alu-46 (358.0 g) followed by BARI Alu-7 (306g), BARI Alu -62 (256.6g), BARI Alu- 36 (194.2g), BARI Alu- 40 (182.4g), BARI Alu-35(142g) and BARI Alu37(113.4g).
IDENTIFICATION OF A NEW CHEMICAL AS A SUBSTITUTE OF AMMONIUM NITRATE (NH 4 NO ) IN MS MEDIA PREPARATION FOR IN VITRO REGENERATION OF POTATO (Solanum tuberosum L.)
(Department of Biotechnology, Sher-e-Bangla Agricultural University, 2016-12) BASHAR, MD. ABUL
An experiment was undertaken at the Biotechnology Laboratory, Department of Biotechnology, Sher-e-Bangla Agricultural University (SAU), Dhaka, Bangladesh during the period of January, 2015 to June, 2016 to identify a new chemical for the substitute of explosive chemical Ammonium nitrate (NH 4 NO ) in MS media composition for in vitro regeneration of potato (Solanum tuberosum L.). Three potato varieties viz Diamant, Cardinal and Asterix were used as an experimental materials. The new chemical was denoted as β chemical for the privacy of the experimental findings. Different concentrations (1, 5. 10, 15 and 20 gmL 3 -1 ) of β chemical were used for the preparation of stock solution- A, which was finally used for MS media preparation. It was noticed that in vitro regeneration of potato has been successfully done with new composition of β chemical in MS media preparation. The shoot length, node number, number of leaf, root length were highest in 5 gmL -1 of β chemical at 14, 21 and 30 days after sub-culture (DAS). Although, in some parameters liquid MS standard dose (16.5 gmL ) gave maximum result but it was statistically non significant with 5 gmL of β chemical. All the plantlets were died at higher concentration (15 gmL and 20 gmL -1 ) of β chemical. It may be due to toxic effect of the chemical. The variety Diamant showed best performance in all the traits under studied. It is reveal that ready made MS powder, Liquid MS standard dose and 1 gmL of β chemical showed comparatively similar result on most of the parameters under studied. Among the varieties, the Asterix variety showed minimal performance in most of the phenotypic characters under studied. In all the case, Diamant variety showed best performance for all the phenotypic characters under studied. The newly used β chemical is cheap, non explosive and environmental friendly which can be an alternative of destructive chemical Ammonium nitrate -1 -1 -1 -1 (NH 4 NO ) for MS media preparation. 3
MICROPROPAGATION OF SALT TOLERANT EXOTIC POTATO GENOTYPES AND THEIR IN VITRO BIOASSAY
(Department of Biotechnology, Sher-e-Bangla Agricultural University, 2016-12) RAHMAN, MD. HABIBUR
In vitro bioassay of nine exotic potato genotypes namely CIP102, CIP106, CIP111, CIP 117, CIP 120, CIP124, CIP127, CIP 136 and CIP 139 was conducted for salinity tolerance at Tissue Culture Lab, Tuber Crops Research Center (TCRC), Bangladesh Agricultural Research Institute (BARI), Gazipur-1701. Single node and root tip segments of these genotypes were cultured in MS media supplemented with 0.0 (control), 80, 100, 120, 140 and 160 mM NaCl. In case of in vitro shoot bioassay, CIP 139 was found as the most salt tolerant with the highest plant height (9.67 cm), number of nodes (9.50), number of leaves (13.60), number of roots (8.00), length of root (6.50 cm), fresh weight of shoot (509.0 mg) and fresh weight of root (205.60 mg) at 160 mM NaCl (14.61 dSm ). On the other hand, CIP 106 was found as the most salinity sensitive at 120 mM NaCl (10.96 dSm -1 ) producing minimum plant height (7.17 cm), number of nodes (6.50), number of leaves (12.50), number of roots (9.70), length of roots (5.10 cm), fresh weight of shoot (572.3 mg) and fresh weight of root (244.4 mg) followed by CIP 136, CIP 117 and CIP 111 at same salinity level. However, CIP 127, CIP 102 and CIP 124 genotypes showed very good performance up to 140 mM NaCl (12.78 dSm -1 -1 ). In vitro root bioassay also revealed the highest salinity tolerance of CIP 139 up to 160 mM NaCl, CIP 127 and CIP 102 up to 140 mM NaCl as well as CIP 106 up to 120 mM salinity level. The root tips of experimented potato genotypes were not significantly affected up to 120 mM salinity level in comparison with control where CIP 120 was found as the lowest tolerant up to 120 mM salinity level in MS media. Among the 0.0 (control), 0.1, 0.5, 1.0 and 1.5 mg /L IBA concentrations supplemented with MS media 1.0 mg/ L was found best for rooting with early root initiation (6.57 days) and well development (10.48 days), highest root length (8.63 cm), 2 highest root number (17.25) and maximum fresh weight of root (332.22 mg). Interestingly, there was no significant differences between 0.5 and 1.0 mg/L IBA for in vitro root induction and development in the experimented CIP potato genotypes.
PHENOTYPIC CHARACTERIZATION AND GENETIC DIVERSITY ANALYSIS OF EGGPLANT (Solanum spp.) GERMPLASM USING SSR MARKER
(Department of Biotechnology, Sher-e-Bangla Agricultural University, 2016-12) TASRIN, KASHPIA
An experiment was carried out with twenty five local and two wild relatives (Solanum sisymbriifolium and S. torvum) of eggplant at the research field and in the Biotechnology Laboratory of Sher-e-Bangla Agricultural University, Dhaka, Bangladesh, during October 2015 to June 2016 to investigate the phenotypic characterization and molecular diversity analysis of different local eggplant germplasm and its wild relatives. The morphological study revealed diversified characters among all the genotypes. Variations were observed in color of leaf vein, stem as well as fruit size and shape. Maximum yield per plant was recorded in Salta Begun and it was 9.8 kg. The minimum yield per plant was observed in both the wild species (3.8 kg) as they produced very small fruits. Correlation of yield contributing traits separated all the genotypes into four clusters (A, B, C, and D). Among them cluster D had five genotypes together with the wild species indicated their relatedness. Five well-known SSR primers were used for the molecular characterization of the genotypes. Ten alleles, ranged from 1 to 3 alleles per locus and an average of 2.0 were detected. Among five primers three were able to produce polymorphic bands and the total number of polymorphic bands was five. The highest (2) number of bands was observed in SSR primers EPSSR82 and smSSR01. The Polymorphism Information Content (PIC) of SSR markers ranged from 0.37 (smSSR01) to 0.67 (EPSSRR82) with an average value of PIC = 0.54. Gene diversity ranges from 0.49 (smSSR01) to 0.72 (EPSSR82), with an average value of 0.61. The value of pair-wise comparisons of Nei‟s (1972) genetic distance (D) and identity between varieties was computed from combined data for the five primers, ranged from 0.20 to 1.00, with an average value of 0.60. Multivariate cluster analysis on the basis of Nei‟s genet ic distance and identity revealed that within those 27 genotypes there surely do exist considerable diversity. UPGMA method separated the segregation of 27 genotypes into two major clusters (I and II). From the clusters, wild species Solanum torvum belonged to the sub-cluster (IIb), that revealed its distinct variation from the others. On the other hand, wild species Solanum sisymbriifolium showed a close relatedness by forming the same cluster together with thirteen local eggplant genotypes. This experiment brought a great opportunity for eggplant improvement as the local genotypes had a close relation with the wild species.
RAPID GENOMIC DNA EXTRACTION IN BULB CROPS AND ESTIMATION OF GENETIC DIVERSITY IN GARLIC (Allium sativumL.) USING RAPD MARKER
(Department of Biotechnology, Sher-e-Bangla Agricultural University, 2016-12) SHEULY, KAMRUN NAHAR
An experiment was carried out in Biotechnology Laboratory of the Department of Biotechnology, Sher-e-Bangla Agricultural University (SAU), Dhaka, Bangladesh to develop low cost and rapid genomic DNA isolation technique and to study the DNA fingerprinting and genetic diversity of nine garlic genotypes. Conventional CTAB/SDS method of DNA extraction required more time and cost. In contrast, the newly developed method to low cost and very fast method for extraction of genomic DNA in bulb crops. The garlic germplasms were BARI-G1, GC001, GC008, GC0012, GC0013, GC0017, GC0024, GC0027 and GC0028. Genomic DNA was extracted in both methods to perform PCR reaction with seven RAPD primers. Sum of 53 distinct and differential amplified DNA bands were generated from PCR amplification. Out of them, 40 bands (75.88%) were polymorphic and 13 bands (24.12%) were monomorphic. The Nei’s genetic identity among 9 garlic genotypes ranged from 0.8053 to 0.9966 and genetic distance ranged from 0.0053 to 0.2166. The UPGMA Dendrogram segregated the 9 garlic genotypes into two main clusters. The first cluster contained 1 genotypes and the second cluster had 8 genotypes. The Dendrogram also indicated that BARI-G1 vs GC0027 varietal pair showed highest Nei’s genetic distance (0.2166) and GC0012 vs GC0017 varietal pair showed lowest genetic distance (0.0053). The study revealed a significant amount of relationship and genetic diversity among the studied 9 garlic genotypes. The RAPD markers were found to be useful tool for molecular characterization and polymorphism study in garlic genotypes.
IN VITRO PROPAGATION OF GARLIC (Allium sativum L.)
(Department of Biotechnology, Sher-e-Bangla Agricultural University, 2016-12) MUBARRAT, FABEEHA
The research work was conducted in the Laboratory of Biotechnology of the Department of Biotechnology, Sher-e-Bangla Agricultural University, from the period of July 2015 to June, 2016 to observe the response of different plant growth regulators on in vitro regeneration of garlic. The disease free, healthy and sterilized basal part of clove of 2-3 cm length were used as explants. Sterilization was done with 70% ethanol, 0.5% HgCl 2 along with few drops of Tween-20. Explants were placed in MS medium supplemented with the Kinetin (KIN), alone and in combination with 2, 4-Dichlorophenoxy acetic acid (2, 4-D). The highest shoot regeneration percentage (100%) was observed by 3.0 mg/L of KIN alone and by wide range of KIN in combination with 2, 4-D (i.e. all combination of 2.5, 3.0 mg/L KIN with 1.0, 1.5, 2.0, 2.5 mg/L 2, 4-D) . The highest shoot length (34.50 cm) was observed after 3 weeks of initiation in 3.0 mg/L KIN treatment. Combined doses of KIN and 2, 4-D showed 1 shoot after 3 weeks of initiation. Number of leaves did not exceed 3 and it was noticed through all combination of 2.0, 2.5, 3.0 mg/L KIN with 1.0, 1.5, 2.0, 2.5 mg/L of 2, 4-D. The highest root induction (100%) was observed by 2.5 mg/L KIN + (1.0, 1.5) mg/L 2, 4-D. It took minimum 3 days to induce root. The average and the highest length of root (3.5 cm and 2.93 cm respectively) was noticed by application of 3 mg/L KIN and 1.0 mg/L 2, 4-D. Maximum number of roots (10.67) were developed by 3.0 mg/L KIN+ 1.5 mg/L 2, 4-D. After transferring the plantlets in the field condition, 86% survival plants were recorded. Finally, in vitro regeneration of garlic was proved to be less time consuming. Moderate doses of hormones showed satisfactory responses in shoot proliferation and root induction which made the tool cost effective as well.
IN VITRO REGENERATION OF ONION (Allium cepa L.) UNDER SALT STRESS CONDITION
(Department of Biotechnology, 2017) PLABON, MD. ASFIQUR RAHMAN
The present research was carried out in Biotechnology Laboratory of the Department of Biotechnology, Sher-e-Bangla Agricultural University, Sher-eBangla Nagar, Dhaka-1207 from the period of September 2016 to July 2017 for in vitro regeneration of onion (Allium cepa L.) under salt stress condition. Three genotypes namely Faridpuri, Taherpuri and Pusa red (Indian) were used as investigated genotypes in this study. Shoot tip segments of these genotypes were cultured in MS (Murashige and skoog, 1962) media having 0.0 (control), 25, 50, 75 and 100 mM NaCl. The shoot tips of experimental onion genotypes were not significantly affected upto 75 mM salinity level in comparison with control treatment. The experiment was conducted at two factorial (variety and Treatment) Completely Randomized Design (CRD) with 3 replications for each treatment. Salinity response on days required for shoot regeneration varied significantly from different salinity condition. The genotype Faridpuri was found maximum tolerant upto 100mM salinity level with 10.60 cm shoot length and 1.43 cm root length having the highest relative shoot and root growth. Besides, Pusa red was found as salinity sensitive genotype showing the lowest 7.03 cm shoot length and 0.60 cm root length at 100 mM NaCl treatment. However, Taherpuri was found tolerant up to 75 mM salinity level with 8.30 cm shoot length and 1.43 cm root length, respectively. The genotypes Faridpuri, Taherpuri and Pusa red (Indian) can be used for further investigation in field condition to evaluate their performance at various salinity levels. Therefore, a convenient in vitro regeneration protocol of onion genotypes under different salinity level has been developed which can be used for screening salinity tolerant variety of onion within a very short period of time.