Establishment And Optimization Of The Chloroplast Transformation System Of Tetraselmis Subcordiformis

Tetraselmis subcordiformis is a common type of unicellular green alga in the oceanwith important application value in the bait feed, new energy development and medicinal,etc. This article is aimed to establish and optimize the Chloroplast Transformation Systemof Tetraselmis subcordiformis, playing a supporting role in promoting the Tetraselmissubcordiformis and related marine algaes basic research and improving the practicalapplication value.We got six gene fragments Prrn, psbA, TpsbA, TpsbC, TrbcL and psbA-5-UTR fromTetraselmis subcordiformis chloroplast genome use of gene cloning method, with greenfluorescent protein gene Gfp as report gene, we constructed five groups of gene expressionvector through genetically modified on pMD-18T and five vectors pRFA, pRFC, pRFL,pFC and pFL were gotten. Besides, we established carriers pRA, pRC, pRL, pC and pLwithout Gfp.Vectors were transferred into Tetraselmis subcordiformis by the method of gene gun andbuilded the chloroplast genetic transformation system. We chose T. subcordiformisbombarded powder as blank and bombarded carriers pRA, pRC, pRL, pC and pL asnegative control. Observing transient expression of green fluorescent protein in thechloroplasts to estimate the expression efficiency of foreign carrier.The results show, under the control of blank experiment and negative test, only vectorpRFA and pFL showed the biological activity, and the expression ability of promoter Prrnand terminator TpsbA in pRFAis better than pFL, containing PpsbA and TrbcL.