Optimization of explants density for tissue culture propagation of banana cv. ‘GRANDE NAINE’

Recent clinical successes in gene therapy applications have intensified interest in using adeno-associated viruses (AAVs) as vectors for therapeutic gene delivery. Although prototypical AAV2 shows robust in vitro transduction of human hepatocyte–derived cell lines, it has not translated into an effective vector for liver-directed gene therapy in vivo. This is consistent with observations made in Fah−/−/Rag2−/−/Il2rg−/− (FRG) mice with humanized livers, showing that AAV2 functions poorly in this xenograft model. Here, we derived naturally hepatotropic AAV capsid sequences from primary human liver samples. We demonstrated that capsid mutations, likely acquired as an unintentional consequence of tissue culture propagation, attenuated the intrinsic human hepatic tropism of natural AAV2 and related human liver AAV isolates. These mutations resulted in amino acid changes that increased binding to heparan sulfate proteoglycan (HSPG), which has been regarded as the primary cellular receptor mediating AAV2 infection of human hepatocytes. Propagation of natural AAV variants in vitro showed tissue culture adaptation with resulting loss of tropism for human hepatocytes. In vivo readaptation of the prototypical AAV2 in FRG mice with a humanized liver resulted in restoration of the intrinsic hepatic tropism of AAV2 through decreased binding to HSPG. Our results challenge the notion that high affinity for HSPG is essential for AAV2 entry into human hepatocytes and suggest that natural AAV capsids of human liver origin are likely to be more effective for liver-targeted gene therapy applications than culture-adapted AAV2.

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Microbulb and plantlet formation of a native bulbous flower, Lilium monodelphum M. Bieb, var. Armenum, through tissue culture propagation

Biotechnology Reports ◽

10.1016/j.btre.2021.e00665 ◽

2021 ◽

pp. e00665

Author(s):

F.N. TOPALOGLU ◽

G. ÖZTÜRK

Keyword(s):

Tissue Culture ◽

Plantlet Formation ◽

Tissue Culture Propagation

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Transmission of Activated-Episomal Banana streak OL (badna)virus (BSOLV) to cv. Williams Banana (Musa sp.) by Three Mealybug Species

Plant Disease ◽

10.1094/pdis-92-8-1158 ◽

2008 ◽

Vol 92 (8) ◽

pp. 1158-1163 ◽

Cited By ~ 29

Author(s):

J. B. Meyer ◽

G. G. F. Kasdorf ◽

L. H. Nel ◽

G. Pietersen

Keyword(s):

Tissue Culture ◽

Enzyme Linked Immunosorbent Assay ◽

Planococcus Citri ◽

Pseudococcus Longispinus ◽

Musa Sp ◽

Transmission Study ◽

Mother Plants ◽

History Of ◽

Tissue Culture Propagation ◽

Transmission Studies

Four different mealybug species (Dysmicoccus brevipes, Planococcus citri, P. ficus, and Pseudococcus longispinus) were evaluated for their ability to transmit putative activated-episomal Banana streak OL (badna)virus (BSOLV) to banana cv. Williams (Cavendish subgroup, AAA). Expressible endogenous sequences of banana streak viruses (BSVs) have been reported to be present in the DNA of various Musa hybrids, including FHIA-21 (AAAB). To obtain activated episomal BSOLV for this experimental transmission study, intentional stress by tissue culture propagation was applied to indexed FHIA-21 which, while free of other viruses, can contain activated episomal BSOLV. Immunocapture polymerase chain reaction and triple-antibody sandwich enzyme-linked immunosorbent assay results revealed that 13.4% of the derived progeny of the mother plants were infected with episomal BSOLV. Four of these BSOLV-infected progeny were used as sources of episomal virus for transmission studies. D. brevipes, Planococcus citri, and P. ficus mealybugs were able to transmit the putative activated episomal BSOLV. Control plants for the transmission experiments included FHIA-21 corms with no background history of tissue culture, as well as virus-free Williams plants. Episomal Banana streak GF (badna)virus (BSGFV) was transmitted from asymptomatic corm-derived FHIA-21 plants by P. citri and P. ficus. This is the first report of P. ficus as a vector of BSVs.

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